Target+-++

YP_004029639.1 1U2P Mycobacterium Tuberculosis is the causative agent of most cases of tuberculosis. Tuberculosis is a common, infectious disease that is oftentimes fatal. It normally attacks the lungs, but can attack other parts of the body too. It's can be spread by an infectious person coughing or sneezing and another person being within the vicinity of the virus in the air. People who have HIV and live in poverty stricken areas are more susceptible to contracting the disease. Most infections don't show sypmtoms, but when they do, they may include a chronic cough, fever, night sweats, and weight loss. Its current main method of prevention is through vaccination and informing people about the risks of the disease.
 * Target (protein/gene name):** tyrosine-phosphatase
 * NCBI Gene # or RefSeq#:** [|FR687359.1]
 * Protein ID (NP or XP #) or Wolbachia#:**
 * Organism (including strain):** Mycobacterium Tuberculosis
 * Etiologic Risk Group (see link below):**
 * Background/Disease Information (sort of like the Intro to your Mini Research Write up):**
 * Essentiality of this protein:** essential
 * Complex of proteins?:** no
 * Druggable Target:** yes

http://www.brenda-enzymes.org/php/result_flat.php4?ecno=3.1.3.48&Suchword=&organism[]=Mycobacterium+tuberculosis&show_tm=0 http://www.sigmaaldrich.com/etc/medialib/docs/Sigma/Datasheet/5/t1196dat.Par.0001.File.tmp/t1196dat.pdf http://www.ncbi.nlm.nih.gov/pubmed/15588980 $61.40 per 1ML Recommended dilution is 1 mL of the cocktail per 500 mg of protein extracted from the tissue or cells. http://www.sigmaaldrich.com/catalog/product/sigma/p0044?lang=en®ion=US
 * *EC#: ** 3.1.3.48
 * Link to BRENDA EC# page:**
 * --** Show screenshot of BRENDA enzyme mechanism schematic
 * Enzyme Assay information (spectrophotometric, coupled assay ?, reagents):**
 * -- link to Sigma (or other company) page for assay or assay reagents (substrates):**
 * -- link (or citation) to paper that contains assay information:**
 * -- List cost and quantity of substrate reagents and supplier:**
 * 1.** p-nitrophenyl phosphate (pNPP) - Phosphatase Inhibitor Cocktail 3DMS

http://www.sigmaaldrich.com/etc/medialib/docs/Sigma/Datasheet/7/f2678dat.Par.0001.File.tmp/f2678dat.pdf
 * 2.** fluorescein diphosphate (FDP)

$208 per 20UG Protein tyrosine phosphatase. (PTP-IA2, ICA-512), Catalytic domain, (amino acids 693- 979), (Gene bank accession no. NM_002846), with N-terminal GST tag, MW = 58.8 kDa, expressed in an //E. coli// expression system. http://www.sigmaaldrich.com/catalog/product/sigma/srp0221?lang=en®ion=US
 * 3.** 6,8-difluoro-4-methylumbellyferyl phosphate (DiFMUP)

-- PDB # or closest PDB entry if using homology model: 1U2P
 * Structure Available (PDB or Homology model)**

It has a high yield for bacterial expression (~1000 mg/L of TB) but the expressed protein was only found in insoluble precipitates. The most often used method for expression is by insect cell culture. The protein is purified under denaturing conditions and yields active kinase.
 * Current Inhibitors:** ALN, orthovanadate, and PAO
 * Expression Information (has it been expressed in bacterial cells):**
 * Purification Method:**

__//**Reference for protein purification and expression:**//__ http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2253236/

MSDPLHVTFVCTGNICRSPMAEKMFAQQLRHRGLGDAVRVTSAGTGNWHVGSCADERAAGVLRAHGYPTD HRAAQVGTEHLAADLLVALDRNHARLLRQLGVEAARVRMLRSFDPRSGTHALDVEDPYYGDHSDFEEVFA VIESALPGLHDWVDERLARNGPS
 * Image of protein (PyMol with features delineated and shown separately):**
 * Amino Acid Sequence (paste as text only - not as screenshot or as 'code'):**


 * length of your protein in Amino Acids:** 163
 * Molecular Weight of your protein in kiloDaltons using the [|Expasy ProtParam] website:** 17.892
 * Molar Extinction coefficient of your protein at 280 nm wavelength:** 15595 M-1 cm-1
 * TMpred graph Image ** (@http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it.



GTGTCTGATCCGCTGCACGTCACATTCGTTTGTACGGGCAACATCTGCCGGTCGCCAATGGCCGAGAAGA TGTTCGCCCAACAGCTTCGCCACCGTGGCCTGGGTGACGCGGTGCGAGTGACCAGTGCGGGCACCGGGAA CTGGCATGTAGGCAGTTGCGCCGACGAGCGGGCGGCCGGGGTGTTGCGAGCCCACGGCTACCCTACCGAC CACCGGGCCGCACAAGTCGGCACCGAACACCTGGCGGCAGACCTGTTGGTGGCCTTGGACCGCAACCACG CTCGGCTGTTGCGGCAGCTCGGCGTCGAAGCCGCCCGGGTACGGATGCTGCGGTCATTCGACCCACGCTC GGGAACCCATGCGCTCGATGTCGAGGATCCCTACTATGGCGATCACTCCGACTTCGAGGAGGTCTTCGCC GTCATCGAATCCGCCCTGCCCGGCCTGCACGACTGGGTCGACGAACGTCTCGCGCGGAACGGACCGAGTT GA
 * CDS Gene Sequence (paste as text only):**

66%
 * GC% Content for gene:**
 * 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) -- 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 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)


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

http://www.brenda-enzymes.org/php/result_flat.php4?ecno=3.1.3.48&Suchword=&organism[]=Mycobacterium+tuberculosis&show_tm=0 http://www.rcsb.org/pdb/explore/explore.do?structureId=1U2P

http://www.sigmaaldrich.com/etc/medialib/docs/Sigma/Datasheet/5/t1196dat.Par.0001.File.tmp/t1196dat.pdf