*Target (protein/gene name): Chorismate Synthase
*NCBI Gene # or RefSeq#: PATRIC ID fig|1121102.3.peg.129
*Protein ID (NP or XP #) or Wolbachia#: No protein ID listed on PATRIC
*Organism (including strain): Campylobacter jejuni
Etiologic Risk Group (see link below): Category B Priority
*/Disease Information (sort of like the Intro to your Mini Research Write up):
Campylobacteriosis is an infectious disease caused by ingesting food or drink contaminated with a bacteria called Campylobacter. Campylobacteriosis is estimated to be the second most common foodborn illness, affecting 1.3 million people each year (according to the CDC). Complications with campylobacteriosis can cause chronic illnesses. For example, people infected with Campylobacter have been known to develop severe arthritis and Guillain-Barré Syndrome (causes acute paralysis and potentially respiratory problems).
Chorismate synthase is part of the shikimate pathway, an essential pathway in C. Jejuni. The shikimate pathway, links metabolism of carbohydrates to the biosynthesis of aromatic compounds. Chorismate synthase catalyzes the conversion of ESPS to chorismate in the last step of the shikimate pathway.
Link to TDR Targets page (if present): No TDR page
Link to Gene Database page (NCBI, EuPath databases -e.g. TryTryp, PlasmoDB, etc - or PATRIC, etc.)
https://www.patricbrc.org/portal/portal/patric/Feature?cType=feature&cId=PATRIC.1121102.3.ARGD01000025.CDS.9220.10320.fwd
Essentiality of this protein: Chorismate synthase has been found to be essential to C. jejuni as it is involved in the synthesis of aromatic amino acids.
Is it a monomer or multimer as biological unit? (make prediction at http://www.ebi.ac.uk/msd-srv/prot_int/pistart.html): Multimer according to the PSA query.
Complex of proteins?: Chorismate synthase is involved in the shikimate pathway, though catalyses the final step independently. It uses flavin mononucleotide (FMN) as a cofactor.
Druggable Target (list number or cite evidence from a paper/database showing druggable in another organism):
http://www.brenda-enzymes.org/enzyme.php?ecno=4.2.3.5
BRENDA shows inhibition of chorismate synthase in other bacteria species.

*EC#: 4.2.3.5
Link to BRENDA EC# page:
http://www.brenda-enzymes.org/enzyme.php?ecno=4.2.3.5
Reaction schematic could not be found on BRENDA; the following schematic is from Wikipedia

ChorismateSynthaseReaction.png

Enzyme Assay information (spectrophotometric, coupled assay ?, reagents):
A spectrophotometric assay of chorismate synthase had been developed. It involves the photoreduction of FMN (flavin mononucleotide) mediated by oxalate. Chorismate formation is monitored at 275 nm. The additions (minus the enzyme) are 50 µM EPSP (substrate), 10 µM FMN (cofactor), 1 mM sodium oxalate, 50 mM phosphate, or bis-Tris propane buffer. Photoreduction was done by illumination by white light (250-W halogen bulb).
Sigma Aldrich has Shikimate-3-phosphate (supposedly used in the enzyme assay in the research paper) for $230.50 for 1 mg. Flavin mononucleotide is available from Sigma Aldrich at $25.00 for 100 mg.
-- link to Sigma (or other company) page for assay (see Sigma links below)
http://www.sigmaaldrich.com/catalog/product/sigma/e0377?lang=en®ion=US (substrate)
-Provided by Sigma; E0377; $114 for 5 mg
http://www.sigmaaldrich.com/catalog/product/aldrich/cds020791?lang=en®ion=US (cofactor)
-Provided by Sigma; CDS020791; $25 for 100 mg
-- -or link (or citation) to paper that contains assay information
http://www.sciencedirect.com/science/article/pii/S0003269784713091

Structure (PDB or Homology model)
-- PDB # or closest PDB entry if using homology model: 1SQ1


Current Inhibitors: No current inhibitors for chorismate synthesis in C. jejuni
Expression Information (has it been expressed in bacterial cells): Has been expressed in E. coli according to BRENDA and research papers.
Purification Method: Purified through centrifugation, fractation with ammonium sulfate, DEAE Sephacel chromatography, cellulose phosphate chromatography, and phenyl sepharose chromatography.

Image of protein (PyMol with features delineated and shown separately):
1SQ1_bio_r_500.jpg

*Amino Acid Sequence (paste as text only - not as screenshot or as 'code'):
MNTFGTRLKFTSFGESHGVAVGCIIDGMPAGVKFDEEFLQNELDKRKGGSKFATPRKESDKAQVLSGVFEGYTTGHPIAI
VVFNENAHSKDYDNLKDLFRPAHADFTYFYKYGIRDHRGGGRSSARESVARVAGGAVAAMLLREFDICVQSGVFGVGTFV
SNLKEEEFDFEFAKKSEIFCLDPKLESDFKNEILNARNSKDSVGAAVFTKVSGMLIGLGEVLYDKLDSKLAHALMGINAV
KAVEIGEGINASKMRGSCNNDALKDGKFLSNHSGGILGGISNGENLILKTYFKPTPSIFAKQESIDKFGNNLKFELKGRH
DPCVGVRGSVVASAMVRLVLADCLLLNASANLNNLKNAYGLKLEHHHHHH
*length of your protein in Amino Acids 370
Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website 40316 kDa.
Molar Extinction coefficient of your protein at 280 nm wavelength: 12295 (M-1)(cm-1)

TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it.

TMPRED.21180.7969.gif


*CDS Gene Sequence (paste as text only):
ttgaatacttttggagtaaaacttagacttacaacttttggagaaagccatggcatagca attggtggcgtgcttgatggttttcctgctggagtaaagatagattttgattttttacaa aatgaacttgataaaagaaaacctagctcaaaatttgctaccaaaagaaaagaaagtgat aaagtagaagttttaagcggagtttttgaagggcttagcacgggaactccaattggtttt attataaaaaacgaagatcaaaaaagcaaagactatggaaatttaaaagatatttttaga ccaggacatgcagactatacatatttttataaatacggtataagagattatagaggtggt ggtagaagtagcgcaagagaaagtgcgataagagttgctggtggggcattttgccaaatg cttttaaatgagtttaaaataagtgttgaaagtggagtttttagtgttggggaagtaaat tttaacgctgattttaacgaggagtttaaaaaagggaatttagactttgaatttgctaaa acttcggaaattttctcactttttaaaaattttgatgaagagtttaaatctgaaatatta aaagccaaaaactctcataatagtgttggagcctcagttgtaactatcataaaaaactcc cccataggacttggagaggttttatatgataaatttgatgcaagattagtagctgctatg atggggataaatgctgtaaaagctgttgaaattggaaatggaataaaatcagccaaaatt tatggcgatgaaaacaatgatgaaatttcaaaaaatggttttttgacaaataattcaggt ggaattttgggtggcattacaaacggcgatgatattttaataaaatcatattttaaacca acaccatcaatttttttagaacaaaatactataaatttaagtggagaagatgaaatttgt aagctcaaaggaaggcatgatccttgcgttggaactagaggaagtgtagttgcaactgct atggcaaggctagttacagctgatatgttactactaaatacaagttcaaatttacaaaat cttaaaaaaatttatggctaa
*GC% Content for gene: 31.97%
*CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol (paste as text only):
*GC% Content for gene (codon optimized):

Do Not Need this info for Spring (but still copy these lines to your Target page for now)
Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~40 nt long primers):
(link to DNA Works output text file - that should be saved in your Google Docs folder after you did the primer design protocol)
-- Ask a mentor, Dr. B, or a fellow researcher -how to link a GDocs file if you are not sure how to.

Primer design results for 'tail' primers (this is just 2 sequences):
**