Dr.+B

See the: @Virtual ToDo List Includes: - Biotin:Avidin docking comparison - ICM vs GOLD vs. VINA - Docking Comparison: Rosie (Rosetta) vs. Drug Discovery at TACC (Vina)

do most inhibitors of metallo enzymes try to displace the metals? Or do they bind with the metals present?

102815 List of other labs that studens may go to:

Walter Fast - Oscar, Brianna, Madeline,

Rhana Gosh - Fin, Alberto Tueme Cruz

Hubreigtse - Avery Wolfe

Finkelstein - Brianna (Xenia) G.

Emily Que: we are making a bunch of probes to bind metalloenzymes for imaging, proteomics, and ‘chemical biology” type applications and would maybe have a place for someone with experience in virtual design of molecules to bind to proteins.

Yan Jessie Zhang - Karan K.

Keatinge-Clay - Luis V.

Lauren Webb: - Ruifei Wang

Deprecated Projects: Wolbachia Targets @SRC Project

052815 Use Rosetta Ligand Docking server with advanced students to compare against GOLD/ICM/VINA http://rosie.rosettacommons.org/ligand_docking/submit

Use PyRosetta or FOLDIt GUI interfaces for Rosetta to do loop modeling/ - maybe homology models.

052215 - Walt won $750 faculty resaerch grant for buying inhibitors of NDM-1 to test. __We have to spend this money in 3 months!__

052115- Target ideas from Luis Luis says - his Burkholderia pseudomallei target is a shunt of the glycoxylate pathway which is itself a shunt to the sugar metabolism pathway He recommends: Isocytrate lyase & malate synthase - are actually both good targets within the glyoxylate pathway metabolize fatty acids instead of sugars Pathogenic bacteria and fungi Good in Tuberculosis and Candida albicans Is there a Isocytrate lysas or malate synthase in Burkholderia ?

051815 - John G to download and print some of these posters - and hang in the lab http://cbc.arizona.edu/njardarson/group/top-pharmaceuticals-poster

2015 Summer students to work on 3-D printing - each makes one 'thing' - teach mentors first

051815 DSF - on NDM-1 using EDTA (Does removing metals destabilize the protein) - Oscar did this - on NDM-1 with inhibitors - Oscar did this with L-captopril - on RAS (does fluorogenic probe destabilize RAS - which may explain RAS protein degradation in cells)

051315 Targets that Walt would like us to do: metallo-beta-lactamase and some serine-beta-lactamase targets "Between a collaborator and I, we have expression plasmids for metal-dependent (NDM-1, IMP-7 (similar to structure of IMP-1), VIM-2) beta lactamases and a serine beta lactamase (KPC-2). And we could get others in each category. They are not currently His-tagged, but could serve as templates for PCR.

I said we can order the Oligo primers with His tag and make them in house - but would be good to have the plasmid (non-HIS) as backups.

IMP-7: (metal-dependent) pET-28b His Tag Sequence attached

VIM-2: (metal-dependent) pET-24a No His Tag

KPC-2: (serine beta lactamase) pET-24a His Tag

ToDo: - sequence verify these: IMP-7: (metal-dependent), pET-28b, His Tag VIM-2: (metal-dependent), pET-24a - No His Tag KPC-2: (serine beta lactamase), pET-24a, His Tag

Re-clone this one into pNIC-Bsa4 to get HIS Tag: VIM-2: (metal-dependent), pET-24a - No His Tag

NOTES ON IMP7 vs. IMP1: We were originally going to go with IMP-1, but Crowder (who sent the plasmid) tells me that IMP1 crashes out of solution a lot, and they have had better luck with IMP-7, so he sent IMP-7 instead. There are structures for IMP1, but not for IMP7. they appear to be about 90% identical. We could dock to IMP-1 (or a homology model of IMP7) and then use IMP7 for the wet lab work. I have not yet mapped where the substitutions occur. If it is a problem, I can always go back to Mike and ask him for IMP-1.

Map where the changes between IMP7 and IMP1 are using PyMol. Make IMP-7 homology model on ICM Dock against IMP1 structure Dock against IMP7 homology structure
 * ToDo:**

ENTER THE LINKS TO YOUR TARGET PAGES HERE: 6-phosphoglucanate dehydrogenase ( //L. monocytogenes //)...................................................Researcher names 6-phosphoglucanate dehydrogenase ( //T. gondii//)...................................................................Researcher names Hypoxanthine phosphoribosyltransferase ( //P. falciparum //)......................................................Researcher names Histidinol-phosphatase (//M. tuberculosis//)...............................................................................Researcher names Glucose-1-phosphatase (//S. enterica//)....................................................................................Researcher names D-alanine ligase (//K. pneumoniae//).........................................................................................Researcher names S-adeosylmethionine syntheses (//L. major//)...........................................................................Researcher names S-adeosylmethionine syntheses (//P. falciparum//)....................................................................Researcher names S-adeosylmethionine syntheses (//T. brucei//)...........................................................................Researcher names S-adeosylmethionine syntheses (//T. gondii//)...........................................................................Researcher names

@TargetF15 - S-adenosylmethionine Synthetase (Mycobacterium tuberuclosis)....................................Bethany R., Ana R. @TargetF15 - D-alanine Ligase (Listeria monocytogenes).....................................................................Simone M. TargetF15 - 6-phosphoglucanate dehydrogenase, decarboxylating, putative (Trypanosoma cruzi)...Diane C. <span style="font-family: Arial,Helvetica,sans-serif;">TargetF15- 6-phosphogluconate dehydrogenase, decarboxylating, putative (Leishmania major)....Mayur P.
 * 0825/16 - Previous Researcher's target page here - we may have a new set of students work on this for Fall 16**

@TargetF15 - S-adenosylmethionine Synthetase (Burkholderia pseudomallei)......................................Matthew N., Justin H.

= Fall 2015 Targets = Primer Design for Springers: (09/21/15) Oligo Primers:

Tail Primers
 * ProtocolPCR_PrimerDesign_OverlapAssemblyVDS_Summ15** in the GDrive/VDSclass/Protocols/__FirstProtocols__


 * ProtocolPCR_PrimerDesignTails_for_pNIC-Bsa4CloningSumm15.doc** in the GDrive/VDSclass/Protocols/__CloningProtocols__

TargetF15- Inosine 5 Monophosphate Dehydrogenase (Bacillus anthracis).......................................Ashley C., Lisa S. TargetF15 - Adenylosuccinate Synthetase (Plasmodium falciparum).................................................Kevin H. TargetF15 - D-alanine Ligase (Staphylococcus aureus)......................................................................Kevin N., Anthony A. @TargetF15 - Protein Tyrosine Phosphatase (Listeria monocytogenes)..................................................Elizabeth K. Krupa S. Sabryna S. TargetF15 - Serine Threonine Protein Phosphatase 1 (Plasmodium falciparum)...................................Kamia R., Jonathan W., Elizabeth M., Neethu G. @TargetF15 - Secreted Phosphatase (Francisella tularensis) ........................................... <span style="background-color: #ffffff; font-family: Arial,Helvetica,sans-serif;">.....................Juliana Z., Samantha V., Mahin D <span style="background-color: #ffffff; font-family: Arial,Helvetica,sans-serif;">TargetF15 - Protein Phosphatase 2A, Putative (Trypanosoma brucei)..............................................Kylie J., Marisa S., Margot F., Cindy N. <span style="background-color: #ffffff; font-family: Arial,Helvetica,sans-serif;">TargetF15- Inorganic Pyrophosphatase (Rickettsia prowazekii)............................................................Ariella S., Brittan M., Andre P.

@http://omictools.com/certain-infectious-and-parasitic-diseases-c1354-p1.html Try TuberQ & Tuberculist
 * Other Databases to look into for targets:**

=**Fall 2014 Targets (Springers)**= @Target - Phosphoserine phosphatase (Vibrio cholerae).............................................John G., Keenan W., Samantha D., Avery W. Mentor: Oscar and Nicole Target - Serine Threonine Phosphatase STP1, (Streptococcus Agalactiae) ...............Jairo V, Saniya H, Stacy T, Daren N Mentor: Kavya and Melissa @Target - 6-phosphogluconate dehydrogenase, decarboxylating, putative (P. falciparum 3D7) .....................Saul H, Lauren M, Carlos Gio O. Mentor: Vicky and Serena Target - Secreted Phosphatase (Mycobacterium tuberculosis)................................. Justin S., Jazmine A., Anh P., Alberto T. Mentor: Priya and Anita @Target-Protein Tyrosine Phosphatase (Listeria monocytogenes)................................Monica L., Tyler T., James F. Mentor: Grace =**Summer 2014 Targets (Vets)**= @Target - Pteridine Reductase 1, (L. Major)................................................................Andrei L., Hailey D. Target - D-alanine:D-alanine Ligase. (M. tuberculosis)...............................................Charina B., Chris T. Target - 6-phosphoglucanate dehydrogenase (P. vivax).............................................Nikki W., Carolyn T.

Target-protein tyrosine phosphatase (Entamoeba histolytica)....................................Andee F. (Vicky G.) - ARISp16?
@Target - Secreted Phosphatase (Francisella tularensis).............................................Cidia G. Target - acetoacetyl-CoA reductase - Burkholderia pseudomallei ................................Luis V., Zain A. @Target - GES-5 carbapenemase (K. pneumoniae) .................................................................................................Xenia G. = Summer 2015 Target Discovery =

With a partner, select a target from the TDR Targets exercise to evaluate. @https://docs.google.com/spreadsheets/d/1etngvvFh0LIAGP13vQcgE9CxWjxjQjuxNNFEVzSMDAs/edit?usp=sharing
 * Part Uno**

Put your names in the NOTES A section. (First name Last Initial e.g. Spongebob S.) Determine why it should be keep in consideration or not If NO If YES If MAYBE
 * write notes in the 'NOTES A' field
 * Color that notes cell RED
 * Then move the line to the 'Trash' sheet
 * write notes in the 'NOTES A' field
 * Color that notes cell GREEN
 * leave the line on that page
 * write notes in the 'NOTES A' field
 * Color that notes cell YELLOW
 * leave the line on that page

Once finished - repeat for another target.

As individuals, select one of the Targets from the Spring 2015 Target Discovery on Wikispaces (see below) Choose one that is NOT yours. Then create a new line in the **Targets_VDSSumm15** @https://docs.google.com/spreadsheets/d/1dymtoVCnFwbX-kOaDWTyWh7KrQ5OZXLw5voZP9xaPb0/edit?usp=sharing Rinse and repeat until we have been through them all.
 * Part Dos**

Otros Objetivos Dr. B selections, etc Luis's recommendations
 * Part Tres**

Step A: finish going through Wiki targets and adding them to Targets_VDSSumm15 Step B: check TDRTargetsExercises for verify hits on Main sheet Step C: Transfer top TDR Targets to Targets_VDSSumm15 Step D: Search and enter info for the below Additional Target into Targets_VDSSumm15

=Additional Targets (from Dr .B)= 6-phosphogluconate dehydrogenase - (in other than //T. brucei// or //P. vivax//) --> //P. falciparum// 3D7, or //Leishmania major// Friedlin strain 1.1.1.49
 * Targets from Prior Years Discovery Sessions that were not used:**

D-alanine:D-alanine ligase (in other than M. tuberculosis) 3-oxoacyl-(acyl-carrier protein) reductase, putative (//P. aeruginosa//) 3-oxoacyl-(acyl-carrier protein) reductase, //L. major// strain Friedlin LmOACP 1.1.1.10

Inosine-5'-monophosphate dehydrogenase EC 1.1.1.205 - not that essential ?

Glutathione reductases in any infectious organism 1.6.4.2 and 1.8.1.7

Thiordoxin Reductase in any infectious organism - P. falciparum 3D7

T. brucei Tb927.3.1240 OG4_10586 protein phosphatase 2A, putative

4EM8 Anaplasma AnphA.00455.a ribose 5-phosphate isomerase B

Pfal008434AAA PFE0730c ribose 5-phosphate epimerase (isomerase), putative yes Pentose phosphate pathway should be essential?

3QH4 Mycobacterium MymaA.00277.c Esterase LipW - PNPA assay

Phosphatase in //T. cruzi// http://tdrtargets.org/targets/view?gene_id=34880

View the highlighted targets in this GDrive sheet: (Seattle Targets) - there are about 5-7 of these SSGCIDstructures_noassocpub_1

= Spring 2015 Target Discovery =

**PART 1:**
find genes for these criteria using the TDR Targets webpage and then enter them onto spreadsheet:
 * TDR Targets Searches Exercise (for practice)**
 * PURPOSE:**
 * **Pick a search, Perform the search, See which targets are found in the list.**
 * **If the target is already on the spreadsheet - __go to the next one that hasn't been entered yet.__**
 * Put your name down in the '**SOURCE / PI**' column for that target and fill in the rest of the information

Phosphatases in T. brucei that have no transmembrane regions, have essentiality evidence, and are druggable at least to 0.5. Phosphatases in T. cruzi that have no transmembrane regions, have essentiality evidence, and are druggable at least to 0.5. Phosphatases in P. falciparum hat have no transmembrane regions, have essentiality evidence, and are druggable at least to 0.5. Phosphatases in P. vivax that have no transmembrane regions, have essentiality evidence, and are druggable at least to 0.5. Phosphatases in T. gondii that have no transmembrane regions, have essentiality evidence, and are druggable at least to 0.5. Phosphatases in M. tuberculosis that have no transmembrane regions, have essentiality evidence, and are druggable at least to 0.5. Phosphatases in L. major that have no transmembrane regions, have essentiality evidence, and are druggable at least to 0.5. Reductase in all (except C. elegans) that have no transmembrane regions, have essentiality evidence, and are druggable at least to 0.6 (be sure to search for reductase and check the box for enzyme)

Spreadsheet for TDR Target @https://docs.google.com/spreadsheets/d/1etngvvFh0LIAGP13vQcgE9CxWjxjQjuxNNFEVzSMDAs/edit?usp=sharing

**PART 2:**

 * SELECTING YOUR TARGET**
 * NOTE:** do this ONLY after you have completed the above TDR Targets exercise.

Follow the rest of the Target Discovery Protocol and create a new Wikispaces page that has all the info you need for a target. (the Wikispaces page will link from here)

=**Spring 2015 Target Selection Pages**= ENTER THE LINKS TO YOUR TARGET PAGES HERE *NOTE; Do not use "/" in your page titles "Target - ................<your name(s)>"

**H**ow To Create a New Target page
Login to Wikispaces, Edit __this__ Targets page with a title that will link to your __new__ page, Type a new line that says :

<span style="font-family: Arial,Helvetica,sans-serif;">TargetSp15 - Disulfide Oxidoreductase (Burkholderia Pseudomallei).....................Researcher: Brittan M.

TargetSp15- Acetylglutamate kinase (Rickettsia rickettsii)...............Researcher: Michael S.

Target Sp15 - Glucose-1-Phosphatase (Salmonella Enterica).....................Researcher: Kamia R.

@TargetSp15- Phosphoglycerate kinase (Trypanosoma brucei)..................Researcher: Margot F.

TargetSp15-BETA-KETOACYL-ACP SYNTHASE III (Plasmodium Falciparum).........................Researcher: Bethany R.

TargetSp15- 2-dehydro-3-deoxyphosphooctonate aldolase (Burkholderia pseudomallei).................Researcher: Ashley C.

<span style="font-family: Arial,Helvetica,sans-serif;">TargetSp15 - 2-acetamido-2-deoxy-alpha-D-glucopyranosyl-(1->6)-phosphatidylinositol de-N-acetylase (Trypanosoma brucei)..... Researcher: Cindy N.

@TargetSp15- Triosephosphate Isomerase (Trypanosoma brucei)............Researchers: Neethu G.

@TargetSp15- Serine threonine-protein phosphatase (Plasmodium falciparum)............ Researchers: Ariella S.

TargetSp15-<span style="font-family: Arial,sans-serif;">hypoxanthine phosphoribosyltransferas (Plasmodium falciparum 3D7)......... Researchers: Lisa S.

TargetSp15- Serine/Threonine-Protein Kinase B PKNB (Mycobacterium tuberculosis).......Researchers: Emily B.

<span style="font-family: Georgia,serif; font-size: 110%;">TargetSp15 - Trehalose-6-Phosphate Phosphatase (Aspergillus fumigatus)................Researchers: Andre P. (this page doesn't exist)

@TargetSp15 - Thioredoxin Reductase (Listeria monocytogenes)...............Researchers: Anthony A.

@TargetSp15 - N-Acetyl-gamma-Glutamyl-Phosphate-Reductase (Mycobacterium tuberculosis)...............Researchers: Mayur P.

@TargetSp15 - Thioredoxin Reductase (Mycobacterium tuberculosis)................... Researchers: Jeffrey X.

TargetSp15 - protein-tyrosine-phosphatase A (ptpA) ................Researchers: Karan K.

TargetSp15 - Isoleucine-tRNA Synthetase, Putative (Toxoplasma gondii)...............Researchers: Bianca E.

@TargetSp15 - Phosphotyrosine protein phosphatase (Mycobacterium tuberculosis)......................Researchers: Simone M.

Target Sp15 - Acetyl-CoA carboxylase (Clostridium botulinum) ...............Researchers: Juliana Z. Target Sp15 - Adenylosuccinate synthetase (P. falciparum)...............Researchers: Elizabeth K

TargetSp15 - Inositol Monophosphatase (Mycobacterium tuberculosis)...........Researcher: Steven T.

TargetSp15 - Fab V Enoyl-ACP Reductase (Yersenia Pestis)..................Researchers: Mahin D.

TargetSp15 -NADH-depemdent Enoyl-(acyl-carrier-protein) reductase (Mycobacterium tuberculosis) ............ Researcher: Krupa S.

@TargetSp15 - Glucosyl-3-phosphoglycerate phosphatase (Mycobacterium tuberculosis)..............Researcher: Sabryna S.

@Target Sp-15- HPPK-DHPS, 2-amino-4-hydroxy-6-hydroxymethyldihydropteridine pyrophosphokinase (Francisella Tularensis) ……..Researcher: Kylie J

TargetSp15 - Histone deacetylase (Plasmodium berghei) ....................................................... Researcher: Jonathan Wang

TargetSp15 - NucleosideTriphosphateDiphosphohydrolase (Toxoplasma gondii)..........................Researcher: Kevin H.

@TargetSp15 - Penicillin-binding protein 2x (Streptococcus Pneumoniae)..................................Researcher: Kevin N.

TargetSp15 - Adenylate Kinase ( Leishmania major) ............................Researcher: Samantha Vargas

@TargetSp 15 - E3 Ubiquitin-protein ligase (Salmonella Typhimurium)..............................................Researcher: Matthew N.

@TargetSp15 - Acetate Kinase (Mycobacterium Paratuberculosis)................................................Researcher: Diane C.

TargetSp15 - CAI-1autoinducer synthase (Vibrio Cholerae).............................................................Researcher: Mina C.

@TargetSp15 - ATP-dependent zinc metalloprotease (Bacillus Anthracis)...................................Researcher: Justin H.

TargetSp15 - Acidocalcisomal exopolyphosphatase (Leishmania major ).....................Researcher - Edward S.

For Example: @TargetSp15 - Enzyme Name (Organism name)........................................................Researchers: Dr. B

TargetSp13 - Fab G 3-ketoacyl-(acyl-carrier-protein) reductase .( Rickettsia prowazeki

i )...................................................................Researchers: Priya P. Oscar V Betty H - - ARISp16?

040115 Pulled information off of target page so that Spring15 studetns can start fresh:

Summer 2014 Target Discovery
Pull best Targets from the
 * 1) TDR Target Discovery Exercise
 * 2) Individual's Target pages
 * 3) The SSGCIDstructures_noassocpub_1 (Seattle Structural Genomics)
 * note: these are ones that have PDB structures but no publications !
 * 1) Past searches from 2013 (see Targets/Archive folder
 * 2) New searches

Small groups discuss, compare their top finds.

Select 5 targets for the group to work on this summer

Do we want to use any ongoing targets? (YopH, FtHap, RpFabG)

Tbrucei 6PG - robust assay

Post info to **TargetSelections_VDSSumm15** on Google Drive

Link to Target Selection Form: Spring 2015 []

Spring 2014 Target Discovery
Note from Dr. B. -When searching for structure for Part 1, see bottom of page 4 of the Target Discovery Protocol; begins with "If **no** Xray crystal structure ..." Be sure to search against the PDB database.

Note: When getting amino acid sequence for BlastP, link out from TDR targets to other database and scroll to the bottom.

**PART 1:**
find genes for these criteria using the TDR Targets webpage and then enter them onto spreadsheet:
 * TDR Targets Searches Exercise (for practice)**
 * PURPOSE:**
 * **Pick a search, Perform the search, See which targets are found in the list.**
 * **If the target is already on the spreadsheet - go to the next one that hasn't been entered yet.**
 * Put your name down in the '**SOURCE / PI**' column for that target and fill in the rest of the information

Phosphatases in T. brucei that have no transmembrane regions, have essentiality evidence, and are druggable at least to 0.5. Phosphatases in T. cruzi that have no transmembrane regions, have essentiality evidence, and are druggable at least to 0.5. Phosphatases in P. falciparum hat have no transmembrane regions, have essentiality evidence, and are druggable at least to 0.5. Phosphatases in P. vivax that have no transmembrane regions, have essentiality evidence, and are druggable at least to 0.5. Phosphatases in T. gondii that have no transmembrane regions, have essentiality evidence, and are druggable at least to 0.5. Phosphatases in M. tuberculosis that have no transmembrane regions, have essentiality evidence, and are druggable at least to 0.5. Phosphatases in L. major that have no transmembrane regions, have essentiality evidence, and are druggable at least to 0.5. Reductase in all (except C. elegans) that have no transmembrane regions, have essentiality evidence, and are druggable at least to 0.6 (be sure to search for reductase and check the box for enzyme)

Spreadsheet for TDR Target @https://docs.google.com/spreadsheets/d/1etngvvFh0LIAGP13vQcgE9CxWjxjQjuxNNFEVzSMDAs/edit?usp=sharing

**PART 2:**

 * SELECTING YOUR TARGET**
 * NOTE:** do this ONLY after you have completed the above TDR Targets exercise.

Follow the rest of the Target Discovery Protocol and create a new Wikispaces page that has all the info you need for a target. (the Wikispaces page will link from here)

=**Spring 2014 Target Selection Pages**= ENTER THE LINKS TO YOUR TARGET PAGES HERE *NOTE; Do not use "/" in your page titles Target - Acidocalcisomal Exopolyphosphatase. Leishmania major.........................................................................Ashly O. Target- PHOSPHOTYROSINE PROTEIN PHOSPHATASE PTPB (Mycobacterium tuberculosis) ............................Ariana Lerma

@Target-serine-threonine protein phosphatase. (P. falciparum 3D7)...........................................................................Lauren McGregor - PDB off Target- protein-tyrosine-phosphotase ywlE .......................................................................................................... Taylor Castillo

Target - Aspartate Aminotransferase. Leishmania major (Friedlin strain) ................................................................James F Target - Trypanosoma brucei, protein tyrosine phosphatase ..................................................................................Researcher: Keenan W. Target- Glutamine Synthetase (Mycobacterium tuberculosis) ............. Researcher: Justin S.

@Target - GES-5 carbapenemase (K. pneumoniae) .................................................................................................Xenia G. Target - Calcium Bound LIPL32 (Pathogenic Leptospira).........................................................................................Sara C. Target - Protein phosphatase-beta, putative (P. vivax)............................................................................................Carolyn Trieu

<span class="wiki_link">Target - Tyrosine-protein phosphatase yopH...........................................................................................................Daren N. @Target - glutathione reductase...............................................................................................................................David Ton

Traget- tyrosine hydroxylase (T. gondii Toxoplasmosis)...........Saul H.

Target - Quiescin sulfhydryl oxidase - Trypanosoma brucei.....................................................................................Tyler Tschirhart

Target - dual specificity phosphatase, catalytic domain-containing protein (Toxoplasma gondii)..................................Anh Pham

Target - UDP-N-acetylglucosamine pyrophosphorylase (Aspergillus fumigatus).........................................................Anna Bozhkova @Target - S-adenosylhomocysteine hydrolase (L. major)............................................................................................Andee F. Target - Probable Adenosylhomocysteinase SAHH (S-Adenosyl-L-Homocysteine Hydrolase) (Adohcyase) (Mycobacterium tuberculosis)......Cidia G. @Target - Tyrosyl-tRNA synthetase (L. major)...............................Researcher: Monica L. Target- DIHYDROLIPOAMIDE DEHYDROGENASE LPD (LIPOAMIDE REDUCTASE (NADH)) (Mycobacterium tuberculosis)....Samantha D.  Target - Probable phosphoribosyl-AMP pyrophosphate HisE (Mycobacterium tuberculosis) ................................................................Sanjna Z.

Target - Island Protein C (IglC) //Francisella tularensis ………...................................................................................//Avery W

Target - Serine-threonine specific protein phosphatase (//Taxoplasma Gondii)// .......................................................Jazmine Alanis (genus spelling) Target - <span style="font-family: 'Times New Roman',Times,serif; font-size: 11pt;">serine-threonine protein phosphatase 2b (catalytic subunit), putative ( <span style="font-family: 'Times New Roman',Times,serif; font-size: 15px;">Plasmodium vivax)...................Saniya Hirani

<span style="font-family: 'Times New Roman',Times,serif; font-size: 15px;">Target - UDP-N-acetylbacillosamine N-acetyltransferase (Campylobacter jejuni)………………………............Charina Benavidez

<span style="font-family: 'Times New Roman',Times,serif; font-size: 15px;">Target - Succinate Dehydrogenase (Flavoprotein subunit) SDHA (Succinic dehydrogenase) (Fumarate reductase) (Fumarate dehydrogena).....Nikki Weber

Target - Malate Dehydrogenase (L. major, Friedlin Strain) ........................................................................................Andrei Loghin

Target- Cyclooxygenase-2 (Francisella tularensis)....................................................................................................John G.

Target - acetoacetyl-CoA reductase - Burkholderia pseudomallei ..............................................................................Luis Valencia

Target - Enoyl-acyl-carrier-protein reductase //Plasmodium falcifarum 3D7//...............................................................Researcher: Stacy Tunggadjaja

@Target - deoxyuridine triphosphatase (Leishmania major)..........................................................................................Alberto T.

++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++ Pulled Target pages - so that students can re-start them on their own:
 * Beginning of Fall 2014**

Were done originally in 2012, then in 2013 Fall, now re-using for 2014 Fall Springers Target - S. agalactiae serine-threonine phosphatase STP1 [NOT CLONED LAST YEAR]+page Researcher: **Brandy C. Caroline C, D'Ondria P** JANICE

Target - Protein Tyrosine Phosphatase (Listeria monocytogenes) -- [NOT CLONED LAST YEAR] - low homology Researcher: **James T, Jesus D, Jiaqi Z** BRANDON Target- phosphoserine phosphatase (Vibrio cholerae) //-(//Created //by// William E.) Researcher: **Dax F.G., Will E., Julia C.** SUMAN


 * Target- Protein phosphatase 2C (Toxoplasma Gondii) ---**(Created by Jensen G.) Researcher:**Jensen G, Imran Z**


 * HERE they are under the 2012 banner:**

2012 Targets
//Targets that were not successfully cloned in 2012// Target-serine-threonine protein phosphatase (Leishmania major)-- Paul, Ling, Rishi **[NOT SUCCESSFULLY CLONED]** Target- Protein Tyrosine Phosphate (Yersinia Enterocolitica).......................................... Ivy, **Stephanie, Jennifer [NOT SUCCESSFULLY CLONED]** Target - S. agalactiae serine-threonine phosphatase STP1 --- Suman, **Brandon, Ruifei [NOT SUCCESSFULLY CLONED]**
 * Target- Protein Phosphatase 2B ................................................................................... ** Divya, **Tom, Sajan** **[NOT SUCCESSFULLY CLONED]**
 * Target - Protein Tyrosine Phosphatase (Listeria monocytogenes) ---** Akhilesh, **Mihir, Shane [NOT SUCCESSFULLY CLONED]**

Spreadsheet Target Selections for Fall 13
As static webpage: As spreadsheet you can edit:

Target - Alpha Carbonic Anhydrase (Helicobacter pylori) ..................................................(Created by Marianna U.) Researcher: **Jessica N, Qasim A, Kelly** DANIEL Target - serine/threonine protein phosphatase catalytic subunit, putative (//T. cruzi//) ...........(Created by Priya) Researcher: ** Grant T, Aditi S.** ANDREW Target -Serine, threonine protein phosphatase 2b (catalytic subunit), putative (P. viva)...........(Created by Feng Gao) Researcher: **Anita V, Ariel C, Gautam** <span style="color: #ff0000; font-family: Calibri,sans-serif; font-size: 11pt;">MAX Target-serine-threonine protein phosphatase (Leishmania major)- [NOT CLONED LAST YEAR] Researcher: **Fin G, Daniel D, Manuel Z** MICHAEL, ALDO
 * Targets that we have Oligos for but have not started cloning:**
 * Targets that were not successfully cloned in 2012 (We have Oligos for some of these - but will order fresh oligos).**

Target- Protein Phosphatase 2B .(//T. brucei//)...................................................... [NOT CLONED LAST YEAR] -page Researcher: **Keely W, Nicole W** URVASHI, DIVYA

Researcher: **Brandy C. Caroline C, D'Ondria P** JANICE

Researcher: **James T, Jesus D, Jiaqi Z** BRANDON Researcher: **Dax F.G., Will E., Julia C.** SUMAN

Researcher:**Jensen G, Imran Z** <span style="color: #ff0000; font-family: Calibri,sans-serif; font-size: 11pt;">MAX

Spreadsheet Target Selections for Summer 13
@https://docs.google.com/spreadsheet/ccc?key=0AvGl3lMyhDsodHRXZGxpZTJjMWl0UXN2STYzcmx2R3c&usp=sharing

Codon Frequency Table for E. coli K-12 (class 2)


 * Links to Individual Target Pages**

"Target - Dihydrofolate Reductase-Thymidylate Synthase (DHFR-TS) [Leishmania Major]Researchers: Tony Garcia, Renée French <span style="color: #ff0000; font-family: Calibri,sans-serif; font-size: 11pt;">JANICE Target - <span style="background-color: #ffffff; font-family: arial,sans,sans-serif; line-height: 1.5;">FabG 3-oxoacyl-(acyl-carrier protein) reductase (P. falciparum) .......................... Researchers: Melissa H. & Jason P. - - ARISp16? DANIEL Target- 3D7, glutathione reductase ......................................................................................(Created by Seo Kelly K.) Researchers: Alyssa K. and Stephanie B.

<span style="color: #ff0000; font-family: Calibri,sans-serif; font-size: 11pt;">BRANDON

=
Target - Serine threonine protein phosphatase PP1 (Plasmodium falciparum)..........................(Created by Anita V.) Researchers: Katherine V., Marianna U., and Ashlee W.===== MICHAEL, ALDO Target - Fab G 3-ketoacyl-(acyl-carrier-protein) reductase .( Rickettsia prowazekii )...................................................................Researchers: Priya P. Oscar V Betty H <span style="color: #ff0000; font-family: Calibri,sans-serif; font-size: 11pt;">URVASHI, DIVYA Target_Serine_threonine_protein_phosphatase5_(Trypanosoma brucei)............................... Researcher: Ramiro R. <span style="color: #ff0000; font-family: Calibri,sans-serif; font-size: 11pt;">ANDREW Target - β-carbonic anhydrases (Mycobacterium tuberculosis) ......................................... (Created by Ashlee W.) Researcher: Nicolet F. <span style="color: #ff0000; font-family: Calibri,sans-serif; font-size: 11pt;">ANDREW

Target- 6-phosphogluconate dehydrogenase (T. brucei) .............................................................(Created by Emily J.) Researchers: Kavya K. and Vicky G. - ARISp16?

<span style="color: #ff0000; font-family: Calibri,sans-serif; font-size: 11pt;">MAX Target - Protein Phosphatase 2A, putative (T. Brucei) ............................................................................Researcher: Jacky E & Kevin E <span style="color: #ff0000; font-family: Calibri,sans-serif; font-size: 11pt;">SUMAN

Target - 3-oxoacyl- (acyl-carrier-protein) reductase (Mycobacterium tuberculosis) ............ (Created by Caroline C) Researcher: Grace & Young - ARISp16?

ANDREW

Target-New Delhi Metallo Beta Lactamase .........................................................(Created by Madeline J.) Researchers: Serena Z., Madeline J., Brendan C. <span style="color: #ff0000; font-family: Calibri,sans-serif; font-size: 15px;">URVASHI, DIVYA

Anaplasma, Babesia, Bartonella, Borrelia, Brucella, Burkholderia, Coccidioides, Cryptosporidium, Ehrlichia, Encephalitozoon, Entamoeba, Giardia, Mycobacterium, Rickettsia, Salmonella
 * Seattle Structural Genomics Targets** - see my Targets folder.

Fall 2013
PseudoKnot Project ProMol Project @Celiac Project

Target Selection For**m**
https://docs.google.com/spreadsheet/viewform?formkey=dEFwQWp6M2lPZFRkcl9HcjBrT1lTOXc6MA#gid=0
 * NOTE:** do this **AFTER** looking at the Targets above

@https://docs.google.com/spreadsheet/pub?key=0AvGl3lMyhDsodDZtbjJSSzVITHNuXzlIOVQwYThWRXc&output=html
 * Target Assignments for Springers**

092213 To see what items were missing from your Target page for your grade, see this spreadsheet (it is also in the GDocs/Mis/Targets folder) @https://docs.google.com/spreadsheet/pub?key=0AvGl3lMyhDsodE1kWEFTc2xsTnRLOGhvdE54RGNaVWc&output=html

@Target-Histidinol Phosphatase (Escherichia coli O157 H7)-(Created by Min S.) - wait on this one. Not sure of essentiality of assay. Target:Inositol phosphate phosphatase sopB (Salmonella enterica subsp. enterica serovar Typhimurium) ... (Created by Alice C.) - wait on this - complex of proteins to be functional. Target - Protein Phosphatase 2C, putative (T. cruzi).........................................................(Created by Selena Ilbeig) - wait on this. Not enough similarity to PDB to maka homology structure Target-MtnX phosphatase (Bacillus Anthracis)..................................//...................................(//Created //by// Brandy C.) - wait. Can't find substrate
 * Interesting Ones - but nor pursued for some reason:**

Target- Protein Phosphatase 1 (T. brucei) -Shivani Bindal - 0.3 Druggability, 80% on 69%

Target - serine-threonine protein phosphatase 2C-(T. gondii)---(Created by Gautam W.) Researcher: ( Was a F13 possibility - but linked to wrong thing so right thing was used)

Target - Putative acid phosphatase (Helicobacter pylori)//............// (Created by Hyun-Young L.) Researcher: none. Was a F13 possibility Target- Protein Tyrosine Phosphate (Yersinia Enterocolitica).......... [NOT CLONED in 2012] - low homology 90% on 28% Researcher: none Was a F13 possibility


 * SUMMER TARGET DISCOVERY REVISITS: **
 * MORNING GROUP**

Ashlee, Alyssa, Anita, Young, Katherine V., Manuel, Melissa, Nicolet, Priya, Ramiro, Vicky, William

//Vittaforma corneae// Target- EcMetAP2-Priya P. (double check assay)

//Trypanosoma brucei// Target: Dihydrofolate reductase - thymidylate synthase (Trypanosoma brucei) --- (created by Melissa Huynh)

//Helicobacter pylori// Target - 1-deoxy-D-xylulose-5-phosphate reductoisomerase  (H. pylori ) ................Hyun-Young L.

// Mycobacterium tuberculosis // Target- Dihydrofolate Reductase M. tuberculosis........................Alyssa K.

Target - Dihyrofolate Reductase (Mycobacterium tuberculosis)............................Ramiro Rodriguez (see Alyssa's page) - join them together

Target - Inositol-1-monophosphatase (Mycobacterium tuberculosis) ...................................Nicolet F.

Target: NADH-dependent Enoyl-Acyl carrier protein reductase (M. tuberculosis)..........Katherine V.

Target-protein tyrosine phosphatase (Entamoeba hystolytica).............................................Vicky G.

Target - Serine threonine protein phosphatase (Plasmodium falciparum) .........................................Anita V.


 * AFTERNOON GROUP**

Tony, Brendan, Jiaqi, Karuna, Renee, Kevin, Kavya, Madeline, Mariana, Serena, Oscar, Jason, Laraib, Serena, Grace, Jacky E,

__ Target- dihydrofolate reductase-thymidylate synthase (Leishmania major) __---Kavya K.

//Chlamydia trachomatis// Target - 5-nucleotidase SurE (Chlamydia trachomatis)---Tony G.

Target-tyrosine phoshatase B (Mycobacterium tuberculosis)// ............................................. //Renee F.

//P. falciparum// Target - dihydroorotate dehydrogenase (malaria)--Kevin E.

//Shigella dysenteriae//
Protein-tyrosine-phosphatase Jiaqi Zhang.

Leishmania major Target - Protein Tyrosine Phosphatase Putative (Leishmania major) - Serena Zadoo

//Klebsiella pneumonia// Target-New Delhi Metallo Beta Lactamase ....................................Madeline J.

Target - HPr Kinase (Mycobacterium pneumoniae).............................................Brendan C.

PRING 2013 VDSers
//Target- NADH-dependent fumaratae reductase (Trypanosome brucei)--Grayson N.//

@Target- Eonyl-(acyl carrier protein) reductase (Burkholderia Pseudomallei 1719b)//--//Grant T.
//Vibrio cholerae//

Target-trypanosoma cruzi, protein tyrosine phosphatase-like protein, putative (t. cruzi)---Nicole W.

Target-Tyrosine-protein phosphatase (bacillus subtilis)--Jessica N.

Target-GDP-mannose pyrophosphorylase(GDP-MP) (Leishmania infantum)..................Keely W.

Target- 3D7, glutathione reductase P. falciparum-Seo Kelly K.

//Dengue Virus 2// Target-Serine Protease Nonstructural Protein 3(NS3)--James T. Target - Non-structural protein 2B- Serine protease subunit NS2B (Dengue Virus 3) ...........Nancy I.

//Salmonella enterica//

// Enterobacter aerogenes // Phosphatidylglycerophosphatase B----Qasim A.

Target-cytosolic malate dehydrogenase (Trypanosoma brucei)Aditi S. Target - Inorganic pyrophosphatase (T. brucei)....................................................Shannon R. Target - serine threonine protein phosphatase 5 (T. brucei) ................ Dax FG

Target -  ................<Khady D.> //Mycobacterium tuberculosis//

Target - 1-DEOXY-D-XYLULOSE 5-PHOSPHATE REDUCTOISOMERASE DXR (Mycobacterium Tuberculosis) ................Imran Zafar //Trypanosoma cruzi// Target -Succinate dehydrogenase flavoprotein, putative (T. cruzi) --Daniel D.

Target - protein kinase, putative (T. cruzi) Alakh R.

//Plasmodium vivax//

Target: Dihydrofolate Reductase (Plasmodium Falciparum 3D7)-Shawn Okpara

Target: Protein tyrosine phosphatase, putative (Plasmodium Vivax)--D'Ondria P. Target: Leishmania major, aldehyde reductase --- Spencer Coronado Target: Leishmania major, trypanothione reductase --- Aaron H. Target: Leishmania major strain Friendlin, phosphatase 2c, putative -Jesus De La O //Bacillus Anthracis//

Salmonella typhimurium Target - Salmonella enterica subsp. enterica serovar Typhimurium (S.typhimurium) ...................................Karinna L.

Target - 3-oxoacyl- (acyl-carrier-protein) reductase (Mycobacterium tuberculosis)..............Caroline C. Target -Dihydrodipicolinate Reductase DAPB (DHPR) M. tuberculosis............ So-Youn P. Target: Thioredoxin reductase (Mycobacterium tuberculosis)---Ariel C. Target- Phosphotyrosine Protein Phosphate (Mycobacterium tuberculosis).........Kelsey C. //​Thermoplasma Acidophilum// Target- trehalose-phosphate phosphatase (Thermoplasma Acidophilum).......................Miguel T.


 * TARGETS THAT WE WILL PASS ON **

Target -enoyl-acyl carrier reductase, Toxoplasma gondii --(created by Grace T.) Target - UDP-N-acetylenolpyruvylglucosamine reductase (Steptococcus Pneumonia)<span style="background-color: #ffffff; color: #222222; font-family: arial,sans-serif; font-size: 13px; line-height: 1.5;">---Brendan C.

__ Target- succinate dehydrogenase (Plasmodium falciparum) __---Anita V. coupled assay (Dr B)

//Campylobacter jejuni// Target- 2-Deoxyuridine 5'-triphosphate nucleotidohydrolase (dUTPase)Manuel Z. --not enough info

@Target - NAD-dependent glyceraldehyde-3-phosphate dehydrogenase (Helicobacter pylori).............William I. (protein complex - 'multifunctional protein')

Summer 2013
no files GDocs: Anita, Kavya, Will, Ramiro, Alyssa, Manuel, Brendan,

Lots: Priya, Vicky, Katherine V., Young, Jacky, Melissa,

Spring 2013
set up 'Teams' - Red, Black, Yellow, etc. Each time is assocaited with a mentor -- this is their point of contact for questions -- they could do peer review within their group

have mentors check their Lab Notebooks before submission.

TestBed:

Fall 2012
PyMol Image Contest

Schedule Fall12 Timeline F12 Timeline Vets F12 (deprecated added stuff to Timeline and split the view in half) Daily Schedule Summ12

Planning
Guest speakers?
 * 1) Profs from ACCbiotech (careers in research),
 * 2) Karishma Kaushik
 * 3) Devinder
 * 4) Grace
 * 5) Lawrence

Cloning vs. expression -- clone new targets (phosphatases only) --

PyMol Refresher VIrtual SCreening refresher ICM tutorial Optional: VINA tutorial - before students can run VINA - have them write one bash script

ToDo List (Dr. B):

 * Homology model
 * Enzyme Assay kinetics lab (phosphatase)
 * Journal Clubs
 * revise Tranformation for Midiprep protocol to require 160 ml of LB for __pNIC-bsa4__growth
 * or just say to use all 4 80 ml tubes in Midiprep preparation

Collaboration project with Supramolecular Sensors (Anslyn/Ghanem) stream -- Eman will run it by Anslyn Issues:
 * 1) expression of the enzyme to give to Supramolecular stream to do phage display binding and sequencing
 * phage display screening only selects binders but gives you no information about binding strength or location of binding (active site vs. allosteric)
 * 1) virtual screening of top cyclic peptides from phage display screening into an enzyme active site
 * benefit of virtual is that we can limit the number of peptides that will need to be synthesized
 * 1) enzyme assays with the best cyclic peptides chosen from
 * what enzyme to use - needs to have large active site to accept a peptide. Maybe DXR (cheaper reagents - big NADH site)
 * however if you target the NADH site - you would have more sideeffects in the patient.
 * how to create 'virtual' peptides - probably on ICM

Virtual Work to be done: (post result to Wiki)
ICM
 * regularization vs Optimization vs Minimization (Cartesian, local, global) - do RMSD differences?
 * Flexible Receptor Docking with 1U72
 * Ensemble docking with various PDB models of 1U72
 * Homology model protocol

Avidin/Biotin Test - downloaded some files to a DDFE folder
 * GOLD
 * ICM
 * VINA

ICM vs. GOLD vs. VINA doc 20ligs vs. DHFR receptor - compare ranking, post on Wiki
 * GOLD
 * ICM
 * VINA

Get Markush library to work in Maestro (since no license in ICM)

Maybe have Christina work on RNA screening validation - see PseudoKnot project page
 * add in molecular dynamics (using Maestro) of ICM and GOLD docked compounds to further refine selections

VINA running on TACC - see 'TACC VINA Work' page - use Alex and Christina

2012 Targets
//Targets that were not successfully cloned in 2012// Paul, Ling, Rishi **[NOT SUCCESSFULLY CLONED]** Ivy, **Stephanie, Jennifer [NOT SUCCESSFULLY CLONED]** Divya, **Tom, Sajan** **[NOT SUCCESSFULLY CLONED]** Suman, **Brandon, Ruifei [NOT SUCCESSFULLY CLONED]** Akhilesh, **Mihir, Shane [NOT SUCCESSFULLY CLONED]**

//Leishmania//

//Wolbachia// endosymbiont of //Brugia malayi (//see Wolbachia Targets page too//)// Target - 1-deoxy-D-xylulose 5-phosphate reductoisomerase [DXR] (Wolbachia) Urvashi Target - FabI enoyl-acyl-carrier-protein reductase NADH (Wolbachia) -- Daniel

//Yersinia// Target - Fab V (Yersinia Pestis) - Max, Michael

//Trypanosoma cruzi//
 * Target - dihydrofolate reductase-thymidylate synthase (T. cruzi)--** Alex, Ling,
 * Target - Glyceraldehyde-3-phosphate dehydrogenase (T. cruzi)---** Andrew, Rishi

//Francisella tularensis//
 * Target -1-deoxy-D-xylulose 5-phosphate reductoisomerase (Francisella Tularensis)--**Janice, Aldo **[NOT SUCCESSFULLY CLONED]**

//Trypanosoma brucei//
 * Target - Glyceraldehyde-3-phosphate dehydrogenase (T. Brucei) .................................**...Larry

//Plasmodium falciparum// Target - 1-deoxy-D-xylulose-5-phosphate reductoisomerase (P. falciparum)-Kaarthik, **Andrew, Alex**

//Staphylococcus agalactiae -// Neonatal Septicemia

//Listeria monocytogenes//

Shigellosis //dysenteriae// <span class="ellipsis_text">Target - B-OCTYLGLUCOSIDE ( Shigellosis<span class="ellipsis_text">) --Jiaqi Zhang

//Candida albicans// Target - Methionine Synthase --- Priya A

2011 Targets
Target-Histidine Phosphatase (Francisella Tularensis) ---Christina, Joey, Target-MPTPB(Mycobacterium Tuberculosis) H37Rv- Joshua Target - Protein-Tyrosine Phosphatase Yersinia pestis YopH (Bubonic Plague) J.C. Target-Mycobacterium Tuberculosis PSTP --Sadhana, Michael Target - Carbonic Anhydrase IX (Homo sapiens) ---Vanna, Keyur Target- Carbonic Anhydrase YadF and cynT(E. coli) Target - PTP1b (Homo sapiens) -VDS Staff


 * Targets not pursued:**

//Francisella tularensis//
 * deprecated - Target -3-oxoacyl-acyl-carrier-protein reductase (Francisella Tularensis)---Jennifer R.**

//Leishmania// Target-dUTP diphosphatase (Leishmania major)- Shane -Requires coupled assay -- Dr. B

Clostridium Perfrigens Target- phospholipase C (C. Perfrigens) Sajan this protein is a toxin

//Staphylococcus aureus// Target- YopH (S. aureus)-Stephanie B --primer design is off, see if you can fix it. Redo - DNA works. --Dr. B Target- Inositol-phosphate phosphatase (S. aureus).........................Brandon N. - essentiality?? -- Dr. B Target - NMN phosphatase, Class B acid phosphatase precursor (Staphylococcus aureus)--Mihir - not uploaded to primers -- Dr.B

Target - fructose-1,6-bisphosphate aldolase (P. falciparum) ....................................Ruifei - Requires coupled assay -- Dr. B

Target - V-type H translocating pyrophosphatase -Tom S. - not uploaded to primers - Dr. B


 * ** New Item ** - each target page should include an image of the TMpred graph (@http://www.ch.embnet.org/software/TMPRED_form.html). In put your amino acid sequence to it.

Wolbachia Targets @https://www.wolbachiawebsite.org/ @http://www.a-wol.net/ Q5GRQ1 - Citrate Synthase (//Wolbachia//) - essentiality? Q5GSP1 2-oxoglutarate dehydrogenase complex (//Wolbachia//) - need to check enzyme assay conditions.- Dr. B Q5GSI8 Succinate dehydrogenase flavoprotein subunit, SdhA (//Wolbachia//) - need to check enzyme assay conditions.- Dr. B Q5GTF8 NADH-quinone oxidoreductase subunit D (//Wolbachia//) Q5GS48 Acetyl-CoA carboxylase, carboxyltransferase component (//Wolbachia//) Q5GS70 Thioredoxin reductase (//Wolbachia//) - Good druggability in P. falciparum - Dr. B Q5GSR3 Delta-aminolevulinic acid dehydratase (ALAD) (//Wolbachia//) - hazardous substrate in assay - Dr. B Transketolase - Druggable, coupled assay though (substrate availability?) Q5GTA4 1-deoxy-D-xylulose 5-phosphate reductoisomerase - Dr. B FabG 3-oxoacyl-[acyl-carrier-protein] reductase EC 1.1.1.100 - called 'Short-chain alcohol dehydrogenase family enzyme' FabI enoyl-[acyl-carrier-protein] reductase NADH EC 1.3.1.9
 * •Wolbachia** .....................................................................(Urvashi, Divya, Daniel, Ramya)

deprecated - Target - 2-oxoglutarate dehydrogenase complex (Wolbachia) --- Daniel deprecated - Target- NADH-quinone oxidoreductase subunit D (Wolbachia)Divya deprecated - Target - Acetyl-CoA carboxylase, carboxyltransferase component wolbachia- Daniel deprecated - Target - Transketolase (Wolbachia) //not used//-Urvashi deprecated - Target - FabG 3-oxoacyl-acyl-carrier-protein reductase (Wolbachia) //not used//--Urvashi deprecated - Target - Citrate Synthase (Wolbachia) //not used// - Ramya deprecated - Target - Thioredoxin reductase (Wolbachia) //not used//--Urvashi

- Do TDR targets search with CRITERIA: phosphatase, Essentiality, Druggability > 0.5. Then manually search for those with good Essentiality, Good match to PDB (>50%), and substrates available (Sigma). -- Dr. B
 * •Chagas //(T.cruzi)//** ......................................................................... (Andrew, Ling, Rishi, Alex)

1-deoxy-D-xylulose 5-phosphate reductoisomerase - none found DHFR? FabI enoyl-[acyl-carrier-protein] reductase NADH EC 1.3.1.9 FabG 3-oxoacyl-[acyl-carrier-protein] reductase EC 1.1.1.100 3-oxoacyl-(acyl-carrier protein) reductase, putative - TDRTargets



--NAD pathway of Francisella tularensis is unique @http://www.bio.utsa.edu/labpages/klose/xzogaj.htm
 * •Francisella tularensis** ......................................................(Aldo, Janice, Jennifer R.)

http://www.sciencedirect.com/science/article/pii/S0304416501001131 http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2223557/?tool=pubmed http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.6.1.13 1-deoxy-D-xylulose 5-phosphate reductoisomerase - yes - Dr. B FabG? 3-oxoacyl-[acyl-carrier-protein] reductase EC 1.1.1.100

FabG? 3-oxoacyl-[acyl-carrier-protein] reductase EC 1.1.1.100 FabI enoyl-[acyl-carrier-protein] reductase NADH EC 1.3.1.9
 * •Yersinia pestis**..................................................................(Michael, Max)

http://www.brenda-enzymes.org/php/result_flat.php4?ecno=3.6.1.23 http://modbase.compbio.ucsf.edu/modbase-cgi/model_details.cgi?queryfile=1338502903_581&searchmode=default&displaymode=moddetail&referer=yes&snpflag=& Try this for Leishmania: **LmjF25.1320** - Dr. B
 * •Leishmania** .....................................................................(Sam, Mihir, Shane )

1-deoxy-D-xylulose 5-phosphate reductoisomerase - none found

Kaarthik - Fen1 **//P. falciparum//**

1-deoxy-D-xylulose-5-phosphate reductoisomerase (DXP reductoisomerase or IspC) EC 1.1.1.267, essential = YES, assayable = YES [|__https://www.ncbi.nlm.nih.gov/gene/812223__] [|DNA Output File]
 * //P. falciparum//** (and others)
 * Gene ID:** 812223

Past target 2011 - Target- Carbonic Anhydrase (Salmonella) --Krishna Past target 2011 - Target - Serine-Threonine Phosophatase 2C From Toxoplasma Gondii--Larry Past target 2011 - Target-Apyrase Phosphotase (Shigella flexneri) deprecated - Target-PARP1 (Homo sapiens)

Targets for pNIC-Bsa4 cloning practice
 * HsCD00041026 ||  || CA7 ||
 * HsCD00002513 ||  || PTP1b ||
 * Zhang ||  || Scp1 ||
 * HsCD00040190 || ASU PSI identifier || CA2 ||

Larry Potential Targets Summer 2012
I used TDR to find some potential targets. (It was the easiest to find essentiality data with)

How does the amino acid sequence BLAST against the PDB? - are there any similar enzymes which have a crystal structure? Be sure to check for any Transmembrane regions (not sure if that is on the Target Discovery Protocol or not)

@http://www.tdrtargets.org/targets/view?gene_id=32911 Unfortunately this one looks to have poor essentiality data. Good - has druggability data - unlike the rest of the TDR ones listed For this one - try to follow the paper link in Assayability and figure out exactly how the assay is done (spectrophotometric?, substrates, etc) They do it "spectrophotometrically taking advantage of the release of protons involved in the reaction", [|Here]. I don't understand how the Michaelis–Menten equation is used, or why there are protons released. --- It is splitting water and putting one proton into solution per reaction. The M-M equations is just for the kinetics. We will get to that later.

Here is a link tot he BioMol Green reagent we can use <span style="background-color: transparent !important; background-position: initial initial !important; background-repeat: initial initial !important; border: none !important; display: inline-block !important; float: none !important; font-family: arial,helvetica,sans-serif !important; font-size: 13.333333015441895px !important; font-style: normal !important; font-variant: normal !important; font-weight: normal !important; height: auto !important; line-height: 26px !important; margin: 0px !important; min-height: 0px !important; min-width: 0px !important; padding: 0px !important; text-decoration: underline !important; vertical-align: baseline !important; width: auto !important;"> phosphate detection - but can it detect pyrophosphate (2 phosphates?) On the product page it says one known <span style="border-bottom: 1px solid; color: #f35b00; cursor: pointer; display: inline-block; height: auto; padding-bottom: 1px; width: auto;">__application__ is "tyrosyl-tRNA synthetase (coupled with pyrophosphatase)" so I'm guessing it can detect pyrophosphatases. It means that they use a coupled assay with a phosphatase to cleave the pyrophosphate into two phosphates so that biomol green can be used. -- If so, then we may have the choice of assay - either the colorimetric pH indicators or this Biomol reagent. --Dr. B

@http://www.tdrtargets.org/targets/view?gene_id=257047 HisE - Looks interesting - how about the assay? It is a Phosphoribosyl-ATP pyrophosphatase, so we can use a pyrophosphate assay, which should be readily available. Is the substrate <span style="background-color: transparent !important; background-position: initial initial !important; background-repeat: initial initial !important; border: none !important; display: inline-block !important; float: none !important; font-family: arial,helvetica,sans-serif !important; font-size: 13.333333015441895px !important; font-style: normal !important; font-variant: normal !important; font-weight: normal !important; height: auto !important; line-height: 26px !important; margin: 0px !important; min-height: 0px !important; min-width: 0px !important; padding: 0px !important; text-decoration: underline !important; vertical-align: baseline !important; width: auto !important;"> ? Not sure about this one, couldn't find it in Sigma. It is a small enzyme too 10kDa - so need to be careful with centrifugal filter MWCO.

@http://www.tdrtargets.org/targets/view?gene_id=255685 looks good too For this one can we just use dUTP as the substrate?

@http://www.tdrtargets.org/targets/view?gene_id=256061 Essentiality is not great -exclude this one

@http://www.tdrtargets.org/targets/view?gene_id=6216 Good too: Same His E as 257047 but for M. tuberculosis instead of M. Leprae - what is the difference between the two strains? M. Leprae <span style="background-color: transparent !important; background-position: initial initial !important; background-repeat: initial initial !important; border: none !important; display: inline-block !important; float: none !important; font-family: arial,helvetica,sans-serif !important; font-size: 13.333333015441895px !important; font-style: normal !important; font-variant: normal !important; font-weight: normal !important; height: auto !important; line-height: 26px !important; margin: 0px !important; min-height: 0px !important; min-width: 0px !important; padding: 0px !important; text-decoration: underline !important; vertical-align: baseline !important; width: auto !important;">. It's very difficult to culture since it lacks many vital genes. M. tuberculosis is also highly aerobic, so it infects the respiratory system while M. Leprae does not need as much oxygen, although is still aerobic. They both have the waxy coating that all mycobacteria have.

@http://www.tdrtargets.org/targets/view?gene_id=5895

Some <span style="background-color: transparent !important; background-position: initial initial !important; background-repeat: initial initial !important; border: none !important; display: inline-block !important; float: none !important; font-family: arial,helvetica,sans-serif !important; font-size: 13.333333015441895px !important; font-style: normal !important; font-variant: normal !important; font-weight: normal !important; height: auto !important; line-height: 26px !important; margin: 0px !important; min-height: 0px !important; min-width: 0px !important; padding: 0px !important; text-decoration: underline !important; vertical-align: baseline !important; width: auto !important;"> I had: What does "probable" mean in the title for the 3rd compound protein? How should I interpret the essentiality data? Some of the essentiality studies are split on essential or not. Does the PDB file need to have a substrate?

-- probable' means they haven't proven its role but based on similarity to other sequences, they think they know what it does. Essentiality data is hit or miss - but just go for ones that at least have some essentiality verified PDB file does not 'need' a substrate - as long as you know where the active site is. - Dr. B

@http://www.rcsb.org/pdb/explore/explore.do?job=summary&pdbId=2i34 Here's one from Pubmed for anthrax, but there's no article on it, and I'm not sure how to figure out essentiality. --- Would be easy assay, just not sure if it is an important enzyme. --Dr. B

Look in the PATRIC database and try to find this. There may be some essentiality data there. Also, search pubmed for more papers on this enzyme to see if there is any essentiality data. - Dr. B

When I search for anthrax in Brenda, a lot of the times it will have the enzyme, but when I click on the link I can't find bacillus anthracis on the list.

--- That is ok - it may not be specifically in there - as long as there are other bacterial versions in there then we are relatively confident. --Dr. B

Here are some for F. tularensis: @http://www.brenda-enzymes.org/literature/lit.php4?e=3.1.3.2&r=666833 It seems that this protein already has known inhibitors -- Also, really similar to Joey and Christina's target -- Dr. B

@http://www.rcsb.org/pdb/explore/explore.do?job=summary&pdbId=3nea No essentiality data I don't think. -- Assay not as simple --Dr. B

Trypanasoma cruzi http://www.tdrtargets.org/targets/view?gene_id=54598 Does the known druggable targets suggest that it is too similar to humans? -- Coupled assay -- Dr. B

http://www.tdrtargets.org/targets/view?gene_id=11394 http://www.pdb.org/pdb/explore/explore.do?structureId=2W7T http://www.jbc.org/content/282/16/11858.long http://www.brenda-enzymes.org/php/result_flat.php4?ecno=6.3.4.2 CTP Synthetase for t. brucei

http://www.tdrtargets.org/survey/view/13

[|/?tab=mo&authuser=0#folders/0B4O2KqKh2q_-U2VjeTQwRzU1cjA] Daily+Schedule+Summ12 [|14] [|24] [|S1368764600901778] [|glyceralde3phosdehydrbaci.pdf] [|DisplayMSDSPage.do] [|result_flat.php4] [|lit.php4] [|lit.php4] [|7932587] [|abstract] [|jm9802620]

SUBSTRATES INFO:
See Wu and Knight paper in the Purchasing/Sigma folder for ino on NADH and NADPH degradation: Conclusions: store NADPH above pH 7.5. Can use Tris + NaOH to do this.

Product Information Sheet: @http://www.sigmaaldrich.com/etc/medialib/docs/Sigma/Product_Information_Sheet/2/n4505pis.Par.0001.File.tmp/n4505pis.pdf <span style="font-family: Arial,sans-serif; font-size: 10pt;">This product is soluble in 0.01 M NaOH (100 mg/ml). <span style="font-family: Arial,sans-serif; font-size: 10pt;">Water alone should not be used to prepare solutions <span style="font-family: Arial,sans-serif; font-size: 10pt;">since it tends to be acidic and would decompose b <span style="font-family: Arial,sans-serif; font-size: 10pt;">-NADH. If solutions must be stored for any length of <span style="font-family: Arial,sans-serif; font-size: 10pt;">time, phosphate buffers should be avoided since they <span style="font-family: Arial,sans-serif; font-size: 10pt;">accelerate the destruction of b <span style="font-family: Arial,sans-serif; font-size: 10pt;">-NADH. <span style="font-family: Arial,sans-serif; font-size: 6.5pt;">6,7 <span style="font-family: Arial,sans-serif; font-size: 10pt;">Trizma Ò <span style="font-family: Arial,sans-serif; font-size: 10pt;">(0.01 M, pH 8.5) and MES buffers are better options. <span style="font-family: Arial,sans-serif; font-size: 10pt;">Since b <span style="font-family: Arial,sans-serif; font-size: 10pt;">-NADH solutions are susceptible to oxidation <span style="font-family: Arial,sans-serif; font-size: 10pt;">even at low temperatures, solutions should be prepared <span style="font-family: Arial,sans-serif; font-size: 10pt;">at concentrations no greater than 5 mM, at a pH of <span style="font-family: Arial,sans-serif; font-size: 10pt;">9–11, and stored at 4 ° <span style="font-family: Arial,sans-serif; font-size: 10pt;">C. <span style="font-family: Arial,sans-serif; font-size: 6.5pt;">6 <span style="font-family: Arial,sans-serif; font-size: 10pt;">The presence of light and <span style="font-family: Arial,sans-serif; font-size: 10pt;">heavy metals can accelerate the oxidation process. <span style="font-family: Arial,sans-serif; font-size: 6.5pt;">1 __<span style="font-family: Arial,sans-serif; font-size: 10pt;">If a __ __<span style="font-family: Arial,sans-serif; font-size: 10pt;">low temperature freezer is available (temperatures at __ __<span style="font-family: Arial,sans-serif; font-size: 10pt;">–40 ° <span style="font-family: Arial,sans-serif; font-size: 10pt;">C or colder), more concentrated solutions can be __ __<span style="font-family: Arial,sans-serif; font-size: 10pt;">prepared and stored for years without any loss of __ __<span style="font-family: Arial,sans-serif; font-size: 10pt;">activity. <span style="font-family: Arial,sans-serif; font-size: 6.5pt;">6 __ b <span style="font-family: Arial,sans-serif; font-size: 10pt;">-NADH should be stored ,desiccated and protected <span style="font-family: Arial,sans-serif; font-size: 10pt;">from light, at –20 ° <span style="font-family: Arial,sans-serif; font-size: 10pt;">C. <span style="font-family: Arial,sans-serif; font-size: 6.5pt;">1 <span style="font-family: Arial,sans-serif; font-size: 10pt;">Solutions should be freshly prepared and used promptly <span style="font-family: Arial,sans-serif; font-size: 10pt;">unless extreme care is taken.
 * NADH** - sigma N4505
 * <span style="font-family: 'Arial,Bold'; font-size: 10pt;">Preparation Instructions **
 * <span style="font-family: 'Arial,Bold'; font-size: 10pt;">Storage/Stability **

N7004 http://www.sigmaaldrich.com/catalog/product/sial/n7004?lang=en®ion=US **Storage/Stability**β-NAD is very hygroscopic and should be stored desiccated.Aqueous solutions between pH 2-6, stored as single - use aliquots at - 70°C, are stable for at least 6 months. Neutral or slightly acidic solutionsare stable at 0°C for at least 2 weeks. Solutions arerapidly degraded upon heating and are very labile inalkaline solutions, especially in the presence ofphosphate, maleate, or carbonate. The rates ofdegradation of solutions at different pH and temperature conditions have been reported. Solutionsare also sensitive to light.
 * B-nad** (nicotinamide adenine dinculeotide) B-NAD+

Also see **Biotek** comparison of NADH vs NADPH for making stocks using Tris, EDTA ph 8 @http://www.biotek.com/resources/docs/NADH_App_Note.pdf

@http://www.sigmaaldrich.com/etc/medialib/docs/Sigma/Product_Information_Sheet/2/n6505pis.Par.0001.File.tmp/n6505pis.pdf b <span style="font-family: Arial,sans-serif; font-size: 10pt;">-NADPH is soluble in 0.01 M sodium hydroxide <span style="font-family: Arial,sans-serif; font-size: 10pt;">(50 mg/ml), yielding a clear, light yellow solution. <span style="font-family: Arial,sans-serif; font-size: 10pt;">It is recommended to store Products N1630, N7505, <span style="font-family: Arial,sans-serif; font-size: 10pt;">and N6505 desiccated at –20 ° <span style="font-family: Arial,sans-serif; font-size: 10pt;">C protected from light. <span style="font-family: Arial,sans-serif; font-size: 10pt;">Product N9910 can be stored at room temperature. <span style="font-family: Arial,sans-serif; font-size: 10pt;">The normal impurities and/or decomposition products <span style="font-family: Arial,sans-serif; font-size: 10pt;">are b <span style="font-family: Arial,sans-serif; font-size: 10pt;">-NADP and monophosphoadenosine 5 ¢ <span style="font-family: Arial,sans-serif; font-size: 10pt;">- <span style="font-family: Arial,sans-serif; font-size: 10pt;">diphosphoribose. <span style="font-family: Arial,sans-serif; font-size: 10pt;">It is recommended to prepare solutions fresh and use <span style="font-family: Arial,sans-serif; font-size: 10pt;">promptly, unless you are sure this is an unnecessary <span style="font-family: Arial,sans-serif; font-size: 10pt;">precaution for your work. However, __it has been__ __<span style="font-family: Arial,sans-serif; font-size: 10pt;">reported that a 0.5 mM solution in 0.02 M NaOH __ __<span style="font-family: Arial,sans-serif; font-size: 10pt;">(pH 12.3) showed no loss of purity in a week at 4 ° <span style="font-family: Arial,sans-serif; font-size: 10pt;">C or __ __ - <span style="font-family: Arial,sans-serif; font-size: 10pt;">85 ° <span style="font-family: Arial,sans-serif; font-size: 10pt;">C, but a 13% loss at –20 ° <span style="font-family: Arial,sans-serif; font-size: 10pt;">C. <span style="font-family: Arial,sans-serif; font-size: 7pt;">3 __
 * NADPH** - sigma N6505
 * <span style="font-family: Arial,sans-serif; font-size: 10pt;">Preparation Instructions **
 * <span style="font-family: Arial,sans-serif; font-size: 10pt;">Storage/Stability **

For NADH and NADPH - the concern is that these are reduced and don't want them to be oxidized. So, want to reduce their exposure to oxygen - hence the NaOH.

Enzo Life Sciences: NADPH is unstable in acid media. Stock solutions should be at pH 8. Soluble in water (50mg/ml) or 0.01M sodium hydroxide.

b-NICOTINAMIDE ADENINE DINUCLEOTIDE PHOSPHATE Sodium Salt Product Number N0505 http://www.sigmaaldrich.com/catalog/product/sigma/n0505?lang=en®ion=US Preparation Instructionsβ-NADP+ is soluble in water at 50mg/ml.It is also soluble in methanol, much less soluble in ethanol andpractically insoluble in ether and ethyl acetate.Aqueous solutions stored as frozen aliquots are stable for at least one year.Repeated freeze thaw cycles are not recommended.Storage/StabilityStore at–20ºC Enzo Life Sciecnefor NADP+ http://www.enzolifesciences.com/ALX-480-003/nadp-.-disodium-salt/ soluble in water to 50mg/ml sesnsitive to alkaline - so store in water that is a little __acidic__.
 * NADP+**

P7877 http://www.sigmaaldrich.com/catalog/product/sigma/p7877?lang=en®ion=US ?? - no info on storage
 * 6-phosphogluconate**

Simga: 28007 $246 http://www.sigmaaldrich.com/catalog/product/sigma/28007?lang=en®ion=US H2O: soluble50 mg/mL, clear, colorless storage life and conditions: ? Sigma Tech Support suggests aliquoting and storing in -80degC
 * Crotonyl coenzyme A trilithium sal**t

Kate K also had C6146 - 5mg **2 Butenoyl CoA Lithium Salt** in her freezer. Is this for Fab I? $232.50 __Yes, this is the same thing.__ For some reason this product number is cheaper

Sigma 13368 solubility: 50 mg/ml in H2O storage life and conditions: ? Sigma Tech Support suggests aliquoting and storing in -80degC
 * 1-Deoxy-D-xylulose-5-phosphate sodium salt (DXP)**

FagG acetoacetyl-CoA + NAPDH --> 3-hydroxybutyryl-CoA + NADP+

acetoacetyl-CoA A1625-5MG - $125 NAPDH N6505-25mg - $87

May be try the reverse reaction:
 * NADP+** Product Number N0505 $79.70 for 100 MG - CHEAP

<span style="font-family: Arial,Helvetica,sans-serif; font-size: 90%;">DL-β-Hydroxybutyryl coenzyme A lithium salt <span style="background-color: #ffffff; font-family: Arial,Helvetica,sans-serif; font-size: 11px; line-height: 1.5;">H0261-10MG $218 - Cheaper thatn AAC

=TARGETS=

Yersinia pestis study - old plague not different from new plague.. http://www.ncbi.nlm.nih.gov/pubmed/21993626

Abstract
Technological advances in DNA recovery and sequencing have drastically expanded the scope of genetic analyses of ancient specimens to the extent that full genomic investigations are now feasible and are quickly becoming standard. This trend has important implications for infectious disease research because genomic data from ancient microbes may help to elucidate mechanisms of pathogen evolution and adaptation for emerging and re-emerging infections. Here we report a reconstructed ancient genome of Yersinia pestis at 30-fold average coverage from Black Death victims securely dated to episodes of pestilence-associated mortality in London, England, 1348-1350. Genetic architecture and phylogenetic analysis indicate that the ancient organism is ancestral to most extant strains and sits very close to the ancestral node of all Y. pestis commonly associated with human infection. Temporal estimates suggest that the Black Death of 1347-1351 was the main historical event responsible for the introduction and widespread dissemination of the ancestor to all currently circulating Y. pestis strains pathogenic to humans, and further indicates that contemporary Y. pestis epidemics have their origins in the medieval era. Comparisons against modern genomes reveal no unique derived positions in the medieval organism, indicating that the perceived increased virulence of the disease during the Black Death may not have been due to bacterial phenotype. These findings support the notion that factors other than microbial genetics, such as environment, vector dynamics and host susceptibility, should be at the forefront of epidemiological discussions regarding emerging Y. pestis infections.

Genes of Interest from Wolbachia
Q5GRQ1 - Citrate Synthase Assay on Sigma - DTNB @http://www.sigmaaldrich.com/catalog/product/sigma/cs0720?lang=en&region=US D8130 - DTNB Sigma cheap B6935 - Assay Buffer - unknown O4126 - Oxaloacetic acid, cheap

Q5GSP1 2-oxoglutarate dehydrogenase complex would need to make all enzymes in complex? NADH based assay @http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.2.4.2 Similar PDB: @http://www.rcsb.org/pdb/explore/explore.do?structureId=2JGD Thiamine dependent? AMDP dependent? Inhibited by Oxaloacetate (in paper fromPDB)
 * S1129** **Sigma** - Succinly coA
 * K3752** Sigma - alpha-Ketoglutaric acid

Q5GSI8 Succinate dehydrogenase flavoprotein subunit, SdhA @http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.3.5.1 Assay not good? @http://jb.oxfordjournals.org/content/145/2/229.long (from Brenda link) uses colorimetric at 600 nm for DCIP 2,4-Dichlorophenolindophenol - was used 2,6-Dichlorophenolindophenol - is available - at Sigma (@http://www.sigmaaldrich.com/catalog/product/fluka/33125?lang=en&region=US) Similar PDB (not the one from BLASTP) - just form text search for SDHA @http://www.rcsb.org/pdb/explore/explore.do?structureId=1NEK
 * W327700** - Sigma Succinate

Q5GTF8 NADH-quinone oxidoreductase subunit D @http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.6.99.5 Assay - 2,6 DCIP similar to Succinate dehyrdogenase PDB Blast @http://www.rcsb.org/pdb/explore/explore.do?structureId=2FUG

Q5GS48 Acetyl-CoA carboxylase, carboxyltransferase component @http://www.brenda-enzymes.info/php/result_flat.php4?ecno=6.4.1.3 @http://www.rcsb.org/pdb/explore/explore.do?structureId=3N6R
 * A2056** **Sigma Acetyl CoA**
 * Free phosphate detection after ATP**

Q5GS70 Thioredoxin reductase @http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.8.1.9 @http://www.rcsb.org/pdb/explore/explore.do?structureId=1TDE
 * T0910 Sigma - Thioredoxin $169.50 /mg**
 * NADPH assay**

Q5GSR3 Delta-aminolevulinic acid dehydratase (ALAD)

@http://www.brenda-enzymes.info/php/result_flat.php4?ecno=4.2.1.24 @http://www.jbc.org/content/277/22/19792.long @http://pubs.acs.org/doi/full/10.1021/bi010656k at A555nm assay 2 step with Ehrlich's reagent and Mercury (!)
 * A3785** **Sigma- cheap**

Other Genes of Interest
//Candida albicans// <span style="font-family: Arial,Helvetica,sans-serif; font-size: 12px;">//Candida albicans// are the most common fungi associated with biofilm-related infections, including urinary tract infections, catheter infections, child middle-ear infections, dental plaque, and endocarditis. Although resistance genes, such as MDR, CDR1, and CDR2, have been identified, research has shown that deletion of these genes does not confer sensitivity to drug therapy in mature biofilms. --Tobudi, //et al. Mycoses//. 2012; 55:199-204

Dr. B Trypanosoma brucei - African sleeping sickness "Treatment requires two drugs, suramin and melarsoprol, that are both very toxic and so rare that they must be directly requested from the World Health Organization" http://www.wired.com/wiredscience/2012/04/exotic-travel-serious/#more-107000

Other potential targets: NAD pathway of Francisella tularensis is unique @http://www.bio.utsa.edu/labpages/klose/xzogaj.htm

The number of people who are dying from illnesses that involve vomiting and diarrhea more than doubled between 1999 and 2007, and most of the increase was due to //Clostridium difficile//. -Wired

//Plasmoidum falciparans// (malaria) //- structure available now 3AU8// @Target - 1-deoxy-D-xylulose-5-phosphate reductoisomerase (good target for many)

//Leishmania major// Target - serine threonine protein phosphatase //Leishmania major//

different from most phosphatases (//Bacillus cereus// or //Bacillus anthrax// (?)) - not sure of essentiality species? essentiality? unique from other phosphatases 3Qy6 PDB
 * YwqE**- bacterial phosphatase

essentiality? - but for virulence //Listeria monocytogenes//
 * LipA** - phosphatase


 * LipA** - phosphatase involved in Virulence of //Listeria monocytogenes -// not sure of essentiality but involved in virulence

//Plasmoidum falciparans// (malaria) essentiality? Ribose5P assay at 280nm cheap substrate
 * ribose 5-phosphate epimerase** (RipA or RipB?)

bacteria or protozoan. Virtual done on DdlB (Kovac paper) but now there are structures of DdlA on PDB. Assay is Malachite green (Biomol), Sigma has substrates
 * DdlA - D-Ala:D-Ala ligase**

Chagas disease (//Trypanasoma cruzi)//

//T. cruzi// (and others) essentiality? use malachite green assay and/or pNPP D-myo-Inositol-1-phosphate (sodium salt) form Cayman Chemical #10007777 $53 for 1 mg
 * inositol-phosphate phosphatase (maybe not good, the Druggable links take you to an alternative for Lithium treaments for manic depression - so may not be a good Essentiality in infectious organisms)**

SOD (Superoxide dismutase) in bacteria is used to prevent oxidative stress induced upon it by the human host immune system. (//H. pylori// has PDB, //M. tuberculosis// has PDB). Also, perhaps [|//Burkholderia// homology]model could be made. Coupled assays, though...

- //Homo sapiens// Carbonic Anhydrase 2

White Nose Bat Disease - Fungal @http://vdsstream.wikispaces.com/page/edit/White+nose+bat+disease

Previous Stuff:
//E. coli// carbonic anhydrase - (**can** or **YadF**) have sequence, assay ok, expression?, essentiality? @http://ecogene.org/geneinfo.php?eg_id=EG12319 forms Synthetic lethal with cynT....

//E. coli// carbonic anhydrase - **cynT** @http://ecogene.org/geneinfo.php?eg_id=EG10176

Human carbonic anhydrase 9 (cancer metastasis?) - different enough from others - there is one homology model and one true structure on PDB

M. tuberculosis - carbonic anhydrase **Rv1284** - essentiality M. tuberculosis - carbonic anhydrase **Rn3588c** **canB** - (yes and no essentiality)

Leishmania, Plasmodium, or Toxoplasmosa DHFR, or DHFR-TS
 * FabI** - enoyl-acyl carrier protein reductase
 * FabG** - 3-oxoacyl-[acyl-carrier-protein] reductase

//Clostridium difficile// toxin - would need protease assay or maybe DSF for cleavage products? @http://www.pdb.org/pdb/explore/explore.do?structureId=3PA8 @http://www.ncbi.nlm.nih.gov/pubmed/21095570

//Plasmodium falciparans// - Malaria ubiquitin carboxyl-terminal hydrolase, Ub-AMC fluorescence assay (essentiality?) [|FTIs] can also be used to inhibit farnesylation in [|parasites] such as trypansoma brucii and [|malaria]. Parasites seem to be more vulnerable to inhibition of Farnesyl transferase than humans are. In some cases this may be because they lack Geranylgeranyltransferase I. Thus it may be possible for the development of antiparastic drugs to 'piggyback' on the development of FTIs for cancer research.

TDR Targets search for any Essentiality and Druggability >= 0.5 - yields 1144 hits TDR Targets search for any Essentiality and Druggability >= 0.8 - yields 324 hits TDR Targets search for any Essentiality and Druggability >= 0.9 - yields 41 hits TDR Targets search for any Essentiality and Druggability >= 1.0 - yields 41 hits (same)

TDR Targets search for 'carbonic' and any Essentiality - yields 6 hits (in tuberculosis, leishmania, etc.) TDR Targets search for 'phosphatase' and any Essentiality - yields 170 hits TDR Targets search for 'phosphatase' and any Essentiality and Druggability >= 0.5 - yields 21 hits

pstP Rv0018c Identified in the membrane fraction of M. tuberculosis H37Rv PHOSPHOSERINE/THREONINE PHOSPHATASE PSTP non essential gene by Himar1-based transposon mutagenesis in H37Rv strain Druggability: 0.2 transmembrane portion though - possible to make truncated version?

//T. cruzi// (and others) inositol-phosphate phosphatase EC 3.1.3.25 PNPP (high Km) or Use the following with Malachite Green detection of free phosphates = D-myo-Inositol-1-phosphate (sodium salt) form Cayman Chemical #10007777 $53 for 1 mg = Pubchem: @http://pubchem.ncbi.nlm.nih.gov/summary/summary.cgi?cid=5288642

TDR Targets: inosine 5'-monophosphate dehydrogenase IMP dehydrogenase substrate: NADPH and =Xanthosine 5′-monophosphate (X1000 discontinued from Sigma)= 1 of 5 for essentiality references

//T. cruzi// - ecto-nucleoside triphosphate diphosphohydrolase (infectivity and virulence)

Furin - human protesase used by Dengue and Anthrax for infectivity. Assay? Bioluminescence dependent on Furin - expensive assay? Side effects? Substrates: with CPA95 as substrate/with N-tert-butoxycarbonyl-Arg-Val-Arg-Arg-methylcoumarin amide as substrate with Ac-Arg-Val-Arg-Arg-4-nitroanilide as substrate

Naegleria fowleri - water born amoeba kills in 3-7 days eating brain

http://the-scientist.com/2011/08/01/make-mine-rare/
 * ===A sampling of rare diseases===

|| South Plainfield, NJ || Cystic fibrosis Duchenne muscular dystrophy Hemophilia || III III II || Cranbury, NJ || Fabry disease Pompe disease Gaucher disease || III I II || South San Francisco, CA || Urea cycle disorders Hepatic encephalopathy || III II || Cheshire, CT || Paroxysmal nocturnal hemoglobinuria Atypical hemolytic uremic syndrome Cold agglutinin disease || Approved II II || Novato, CA || Lysosomal storage disorders Phenylketonuria Lambert-Eaton myasthenic syndrome || 2 products approved Approved Approved in EU ||
 * **Company** || **Indication** || **Phase** ||
 * **PTC Therapeutics**
 * **Amicus Therapeutics**
 * **Hyperion Therapeutics**
 * **Alexion Pharmaceuticals**
 * **BioMarin Pharmaceutical**

08/17/2016 Staphylopine as a drug target for bacteria (specifically S. aureus, Y. pestis, and P aeruginosa - one would target the machinery (proteins) that make this small molecule metallophore (e.g. it binds up metals and helps them get transported into the cell so it can survive) There are 3 enzymes supposedly which make this from amino acid lysine. There are also some proteing involved in trafficking staphylopine (move it in and out of cell membrane) - see C&EN news May 30, 2016 edition page 9. - see Pascal Arnoux Bioscience & Biotechnology Institute in Aix-Marseille, France - see Eric Skaar lab of Vanderbilt University.