N-acetylneuraminic+acid+synthase+(Neisseria+meningitidis)

Meningitis is the inflammation of the meninges, the thin layer of membranes that covers the brain and the spinal cord. There are three types of infections: bacterial, viral, and fungal. The most contagious and lethal form is bacterial meningitis; death can occur in as little as a few hours. In the United States, approximately 4,100 cases of bacterial meningitis, including 500 deaths, occurred annually from 2003-2007. Meningitis symptoms include sudden onset of fever, headache, and stiff ache. Additional symptoms include nausea, vomiting, increased sensitivity to light, and confusion. From 1/3 to 1/2 of the survivors face permanent physical or mental damages. Neisseria meningitidis (a gram-negative, aerobic, non-endospore forming diplococcus) is one of several bacteria responsible for the crippling disease. It can be spread from person to person through coughing and sneezing. Inside the nasal cavities, the bacteria quickly divide to enter the bloodstream. Once the bacteria enter the subarachnoid space, the host’s immune system is unable to suppress the infection from developing in the cerebrospinal fluid because the region lacks the necessary antibodies and white blood cells. The pathogens produce sialylated capsular polysaccharides, mimicking the exterior surface of mammalian cells and consequently concealing the bacteria from the host’s immune system One potential inhibitor discovered in la http://pubs.acs.org/doi/abs/10.1021/bi9012758. Also, Brenda shows inhibitors for the target in other organisms. Show screenshot of BRENDA enzyme mechanism schematic 
 * Target (protein/gene name): ** N-acetylneuraminic acid synthase
 * NCBI Gene # or RefSeq #: **902174
 * Protein ID (NP or XP #) or Wolbachia#: **NP_273133.1
 * Organism (including strain): ** //Neisseria meningitidis// MC58
 * Etiologic Risk Group: **2
 * Disease Information: **
 * Link to TDR Targets page (if present): **<span style="font-family: 'Times New Roman',serif;">N/A
 * <span style="font-family: 'Times New Roman',serif;">Link to Gene Database page (NCBI, EuPath databases, -e.g. TryTryp, PlasmoDB, etc –or PATRIC, etc.) **<span style="font-family: 'Times New Roman',serif;">[] (The site titles the gene as “polysialic acid capsule biosynthesis protein SiaC” which is another name for N-acetylneuraminic acid synthase)
 * <span style="font-family: 'Times New Roman',serif;">Essentiality of the Protein: **
 * <span style="font-family: 'Times New Roman',serif;">Is it a monomer or multimer as biological unit? **<span style="font-family: 'Times New Roman',serif;"> Dimer
 * <span style="font-family: 'Times New Roman',serif;">Complex of proteins: ** N/A
 * <span style="font-family: 'Times New Roman',serif;">Druggable target (list number or cite evidence from a paper/database showing draggle in another organism): **
 * <span style="font-family: 'Times New Roman',serif;">EC#: **<span style="font-family: 'Times New Roman',serif;">2.5.1.56
 * <span style="font-family: 'Times New Roman',serif;">Link to BRENDA EC# page: **<span style="font-family: 'Times New Roman',serif;">[]

<span style="font-family: 'Times New Roman',serif;">Continuous spectrophotometric assay <span style="font-family: 'Times New Roman',serif;">Continuous coupled assay for phosphate <span style="font-family: 'Times New Roman',serif; line-height: 1.5;">[] <span style="font-family: 'Times New Roman',serif; line-height: 1.5;">[]
 * <span style="font-family: 'Times New Roman',serif;">Enzyme Assay Information (spectrophotometric, coupled assay, reagents): **
 * <span style="font-family: 'Times New Roman',serif;">Link to Sigma (or other company) page for assay o ****<span style="font-family: 'Times New Roman',serif; line-height: 1.5;">r link (or citation) to paper that contains assay information: **
 * <span style="font-family: 'Times New Roman',serif;">List cost & quantity of substrate reagents, suppliers, & Catalogs #: **
 * <span style="font-family: Times New Roman,serif;">Phosphoenolpyruvate (Sigma Aldrich #10108294001): $255.00
 * <span style="font-family: Times New Roman,serif;">N-acetyl-D-mannosamine (Sigma Aldrich #A8176-10MG): $19.10

<span style="font-family: 'Times New Roman',serif;">- 5-acetamido-4,6,7,8,9-pentahydroxy-2-phosphoryl nonanoic acid triethylammonium salt <span style="font-family: 'Times New Roman',serif;">Inhibitors for the target in other organisms are found on Brenda. http://www.brenda-enzymes.org/enzyme.php?ecno=2.5.1.56&Suchword=&organism%5B%5D=Neisseria+meningitidis&show_tm=0
 * <span style="font-family: 'Times New Roman',serif;">Structure (PDB or Homology Model): **<span style="font-family: 'Times New Roman',Times,serif; line-height: 1.5;">1XUU, 1XUZ
 * <span style="font-family: 'Times New Roman',serif;">Current inhibitors: **

<span style="font-family: 'Times New Roman',serif;">- Has been expressed in E. Coli BL21(DE3) ([]) ([]) <span style="font-family: 'Times New Roman',serif;">- Found in the cytoplasm <span style="font-family: 'Times New Roman',serif;">- Ni2+ Chromatography
 * <span style="font-family: 'Times New Roman',serif;">Expression Information (has it been expressed in bacterial cells): **
 * <span style="font-family: 'Times New Roman',serif;">Purification Method: **
 * <span style="font-family: 'Times New Roman',serif;">Image of Protein (PyMol with features delineated & shown separately): **

<span style="font-family: 'Times New Roman',serif;"> Amino Acid Sequence: >1XUZ:A|PDBID|CHAIN|SEQUENCE: MQNNNEFKIGNRSVGYNHEPLIICEIGINHEGSLKTAFEMVDAAYNAGAEVVKHQTHIVEDEMSDEAKQVIPGNADVSIY EIMERCALNEEDEIKLKEYVESKGMIFISTPFSRAAALRLQRMDIPAYKIGSGECNNYPLIKLVASFGKPIILSTGMNSI ESIKKSVEIIREAGVPYALLHCTNIYPTPYEDVRLGGMNDLSEAFPDAIIGLSDHTLDNYACLGAVALGGSILERHFTDR MDRPGPDIVCSMNPDTFKELKQGAHALKLARGGKKDTIIAGEKPTKDFAFASVVADKDIKKGELLSGDNLWVKRPGNGDF SVNEYETLFGKVAACNIRKGAQIKKTDIE ATGCAAAACAACAACGAATTTAAAATTGGTAATCGTTCAGTAGGTTACAACCACGAACCATTGATTATCT GTGAAATCGGCATCAATCATGAAGGCTCTTTAAAAACAGCTTTTGAAATGGTTGATGCTGCCTATAATGC AGGCGCTGAAGTTGTTAAACATCAAACACACATCGTTGAAGACGAAATGTCTGATGAGGCCAAACAAGTC ATTCCAGGCAATGCAGATGTCTCTATTTATGAAATTATGGAACGTTGCGCCCTGAATGAAGAAGATGAGA TTAAATTAAAAGAATACGTAGAGAGTAAGGGTATGATTTTTATCAGTACTCCTTTCTCTCGTGCAGCTGC TTTACGATTACAACGTATGGATATTCCAGCATATAAAATCGGCTCTGGCGAATGTAATAACTACCCATTA ATTAAACTGGTGGCCTCTTTTGGTAAGCCTATTATTCTCTCTACCGGCATGAATTCTATTGAAAGCATCA AAAAGTCGGTAGAAATTATTCGAGAAGCAGGGGTACCTTATGCTTTGCTTCACTGTACCAACATCTACCC AACCCCTTACGAAGATGTTCGATTGGGTGGTATGAACGATTTATCTGAAGCCTTTCCAGACGCAATCATT GGCCTGTCTGACCATACCTTAGATAACTATGCTTGCTTAGGAGCAGTAGCTTTAGGCGGTTCGATTTTAG AGCGTCACTTTACTGACCGCATGGATCGCCCAGGTCCGGATATTGTATGCTCTATGAATCCGGATACTTT TAAAGAGCTCAAGCAAGGCGCTCATGCTTTAAAATTGGCACGCGGCGGCAAAAAAGACACGATTATCGCG GGAGAAAAGCCAACTAAAGATTTCGCCTTTGCATCTGTCGTAGCAGATAAAGACATTAAAAAAGGAGAAC TGTTGTCCGGAGATAACCTATGGGTTAAACGCCCAGGCAATGGAGACTTCAGCGTCAACGAATATGAAAC ATTATTTGGTAAGGTCGCTGCTTGCAATATTCGCAAAGGTGCTCAAATCAAAAAAACTGATATTGAATAA ATGGAAAAAGTTAATGAAGAACGTGACGCGGTGTTTGAAGATCACATCGGCGACCGTCGCCGCTCTGTTCGTTCTCTGCTCGAAGAAGCTTTCGCGGACGAAATGGAGAAAACCTCTTACGACGTTGAGGTTGCGGATACCCCACAGCCGCACATCCCAATCCGTTTTCGCCACCCGCCGATCGCTGGCCCAGTTCATGATGTGTTCGGTGATGCTATCCACGACATCTTTCAGAAAATGATGAAACGCGGTCAGGCTGTTGACTTCTGCCACTGGGTGTCTCACCTGATTGCGACTGAAATCGACGAAAAGTTTTCCGAAGTTGCGTTCCGTGACGTTCAGTACAACCCGGACATCTACGTTACGGACTCTACTACCGAAGCGAAAAAACTGTTCAACGATAAGATTTGGCCTGCGATCGATAAAATCCTGCAGCAAAACGCCGAAACCTGTCCGATCCTGTCTGAGAAATGGTCTGGTATTCACGTTAGCGGTGATCAGCTCAAGGGTCAGCGCCACAAACAGGAAGATCGCTTCCTGGCTTACCCTAACGGTCAATATATGGACCGTGGTGAGGACCCGATTTCTGTTCTGGCGGTTTTCGATGGTCACGGCGGTCACGAATGCTCTCAGTACGCGGCAGGTCATCTGTGGGAAACGTGGCTGGAGGTGCGTAAAAGCCGCGACCCGTCTGACTCTCTGGAGGACCAGCTGCGCAAATCTCTCGAACTGCTGGACGAACGTATGACCGTGCGTTCTGTCAAAGAATGCTGGAAAGGCGGTTCTACCGCTGTCTGCTGTGCAATCGACATGGACCAGAAGCTCATGGCGCTCGCATGGCTGGGCGACTCTCCGGGCTATGTTATGTCTAACATCGAATTCCGTCAACTCACTCGTGGTCATTCTCCTTCTGATGAGCGCGAAGCACGTCGCGTAGAGGAGGCAGGTGGCCAACTCTTTGTAATCGGTGGTGAACTCCGTGTTAACGGTGTGCTGAACCTGACGCGCGCACTCGGCGATGTTCCGGGTCGTCCGATGATCTCTAACGAACCGGAAACTTGCCAGGTTCCGATCGAATCTAGCGACTATCTCGTGCTCCTGGCGTGCGATGGTATCAGCGACGTATTCAATGAGCGTGACCTGTATCAACTGGTCGAAGCGTTTGCCAATGATTACCCGGTTGAAGACTACGCGGAACTGTCTCGCTTTATTTGCACCAAGGCCATTGAAGCAGGCTCTGCGGACAATGTATCTGTTGTTATTGGTTTCCTGCGTCCGCCGCAGGACGTGTGGAAACTGATGAAGCATGAATCTGACGACGAAGACTCTGATGTTACCGATGAGGAATAA
 * <span style="font-family: 'Times New Roman',serif;">Length of your proteins in Amino Acids **<span style="font-family: 'Times New Roman',serif;">: 349
 * <span style="font-family: 'Times New Roman',serif;">Molecular weight of your proteins in kiloDaltons using the [|Expasy ProtParam Website]: **<span style="font-family: 'Times New Roman',serif;"> 38390.7kDa
 * <span style="font-family: 'Times New Roman',serif;">Molar Extinction Coefficient of your protein at 280nm wavelength: **<span style="font-family: 'Times New Roman',serif;">21890
 * <span style="font-family: 'Times New Roman',serif;">TMpred graph Image (@http://www.ch.embnet.org/software/TMPRED_form.html) **<span style="font-family: 'Times New Roman',serif;"> Input your amino acid sequence to it
 * <span style="font-family: 'Times New Roman',serif;">CDS Gene Sequence: **
 * <span style="font-family: 'Times New Roman',serif; line-height: 1.5;">GC% Content for gene: **<span style="font-family: 'Times New Roman',serif; line-height: 1.5;">40.762%
 * <span style="font-family: 'Times New Roman',serif;">CDS Gene Sequence (codon optimized) copy from output of Primer Design Protocol: **


 * <span style="font-family: 'Times New Roman',serif;">GC% Content for gene (codon optimized **<span style="font-family: 'Times New Roman',serif;">): 52.59%


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

Forward Primer: (Underlined = pLIC Primer Attachment) 5’-- __TACTTCCAATCC__ATGGAAAAAGTTAATGAAGAA --3’ Reverse Primer: (underlined = pLIC Reverse Primer Attachment) 5’-- __TATCCACCTTTACTG__TTATTCCTCATCGGTAACATC --3’