Mycobacterium tuberculosis molecules by Dr. PI

The gene Name is deduced from Isoniazide (INH) which is a first-line antibiotic for the treatment of tuberculosis. Isoniazid is known to inhibit mycolic acid biosynthesis, mainly by preventing the chain extension of long fatty acyl substrates. This suggests that the target of its action is within the mycobacterial FAS-II system.

isoniazide

Enoyl-ACP-reductases are involved in fatty acid biosynthesis. They utilize NADH to reduce the fatty acyl chain linked to the acyl carrier protein. Mycobacteria possess two systems to synthesize fatty acids; FAS-I for fatty acyl chains up to 16 carbons and FAS-II for the production of long chains with up to 56 carbons which are precursors of the mycolic acids, a major component of mycobacterial cell walls. Enoyl-ACP-reductase catalyses the reduction of the double bond between C2 and C3 of the ACP-bound enoylsubstrate, which is the last step in the cyclic elongation of fatty acids.

Enoyl-ACP-reductae reaction

The figure shows the enoyl-[ACP]-Reductase reaction. The atoms shown in red are deduced from Acetyl-CoA that was added in the last elongation cycle of the fatty acid biosynthesis, the .

The from Mycobacterium tuberculosis is a member of the short chain dehydrogenase/reductase (SDR) . belongs to the functional group of genes involved in the synthesis of fatty acids and mycolic acids as shown below. has activity of an oxidoreductase acting on the CH-CH group of donors with NAD⁺ as acceptor.

This molecular model shows named and Each unit consists of a with a central core that contains an NADH binding site. Each subunit contains one molecule and four of the six subunits have bound a in addition. In aequous solution InhA is a with high symmetry and close intramolecular contacts.

Taking a at a single subunit reveals the conformation of the bound substrate and the shape of the binding crevice. The C₁₆ fatty acid acyl substrate folds into a conformation, with the trans double bond located directly over the of NAD⁺. The fatty acid substrate binding crevice of InhA has a oval-shaped cavity with and exposed to the solvent, where the substrate would be attached to the acyl carrier protein. The C2=C3 trans double bond faces the closed end of the crevice and is completely by hydrophobic residues. Additional surround the fatty acyl chain portion of the substrate. Within the NAD binding site the side chain hydroxyl of forms a direct hydrogen bond to the thioester carbonyl oxygen of the substrate. Mutation of can be a cause of INH resistance.

spin / | | 3D model needs Jmol

Isoniazid is a precursor of the antituberculous drug and must be converted into an activated form. This is performed by the mycobacterial catalase-peroxidase (KatG). The activated form of isoniazid (probably an isonicotinic-acyl radical) becomes covalently attached to the nicotinamide ring of the NADH bound within the active site of InhA. This creates an adduct that acts as a very tightly bound inhibitor of the InhA enzymatic activity and thus prevents the synthesis of mycolic acids.

The structural model was obtained from PDB entry , and the description from . Click here to search for at PDB.

is a peptide of with a and a It's position on a has been identified. Using a genetic approach a single open reading frame was identified (inhA), whose amino acid sequence is shown here. Three features are highlighted in the sequence: Mutation in position 94 from serin to alanin is responsible for INH-resistance in Mycobacterium smegmatis by decreasing the affinity for NADH. Region 136-165 is the predicted NAD binding site with the substrate binding at tyrosin 158. The sequence contains several .

has significant aminoacid sequence homology to many other genes within mainly to dehydrogenases and reductases that are involed in steroid and fatty acid metabolism. Comparison to proteins of reveals InhA homologues in other mycobacteria, corynebacteria, brevibacterium and many more.

The gene is encoded in a In the M. tuberculosis laboratory strain H37Rv it is a known as which corresponds to gene in the clinical isolate CDC 1551. It can be found on .

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On the circular M. tuberculosis chromosome lies at and is flanked by the genes fabG1 and hemZ, which are also involved in lipid biosynthesis.

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blueTB molecules are published by Paul Imboden, Dr. PI Bioconsulting. Authorization to photocopy or reproduce this entry for personal use is granted. Copyright @ 2005 Paul Imboden, Dr. PI Bioconsulting. Last modified May 17, 2008 . Disclaimer: blueTB and the author reserves the right to modify and cancel any statement in these documents and regrets, that he cannot accept any responsibility for the consequences of any such changes. to the best of my knowledge all information is correct, but I cannot accept liability for any errors. References for this blueTB entry are:

Deciphering the biology of Mycobacterium tuberculosis from the complete genome sequence. Cole ST, Brosch R, Parkhill J, Garnier T, Churcher C, Harris D, Gordon SV, Eiglmeier K, Gas S, Barry CE 3rd, Tekaia F, Badcock K, Basham D, Brown D, Chillingworth T, Connor R, Davies R, Devlin K, Feltwell T, Gentles S, Hamlin N, Holroyd S, Hornsby T, Jagels K, Barrell BG, et al. Nature, 393:537-44 (1998) and the Mycobacterium tuberculosis sequencing project from the Sanger Centre.

Whole-genome comparison of Mycobacterium tuberculosis clinical and laboratory strains. Fleischmann RD, Alland D, Eisen JA, Carpenter L, White O, Peterson J, DeBoy R, Dodson R, Gwinn M, Haft D, Hickey E, Kolonay JF, Nelson WC, Umayam LA, Ermolaeva M, Salzberg SL, Delcher A, Utterback T, Weidman J, Khouri H, Gill J, Mikula A, Bishai W, Jacobs Jr WR Jr, Venter JC, Fraser CM. J Bacteriol. 184:5479-90 (2002) and the Mycobacterium tuberculosis sequencing project from TIGR .

3. Dr. PI's Mtbook, The Mycobacterium tuberculosis genome in a book. Paul Imboden, Dr.PI Bioconsulting. mtbook.drpi.ch/ Release 1.8.0 Jan 2004, which itself is based mainly on reference 1.

Crystal Structure of the Mycobacterium tuberculosis enoyl-ACP reductase, InhA, in complex with NAD⁺ and a C₁₆ fatty acyl substrate. Rozwarski DA, Vilcheze C, Sugantino M, Bittman R, Sacchettini JC. J. Biol.Chem. 274:15582-1558 (1999).

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