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Category: Microbial Genetics and Molecular Biology
The Molecular Genetics of Mycolic Acid Biosynthesis, Page 1 of 2
< Previous page | Next page > /docserver/preview/fulltext/10.1128/9781555818845/9781555818838_Chap29-1.gif /docserver/preview/fulltext/10.1128/9781555818845/9781555818838_Chap29-2.gifAbstract:
The mycobacterial cell wall is essential for Mycobacterium tuberculosis growth and survival. It is lipid-rich and highly impermeable and thereby provides protection from many antibiotics, and also allows the pathogen to proliferate within macrophages and to persist for extended periods of time in the infected host ( 1 ). Mycolic acids, which are long-chain α-alkyl β-hydroxy fatty acids, constitute up to 60% of the cell wall and are principally responsible for the low permeability of the waxy cell envelope ( 2 ). They are found primarily as esters of the nonreducing arabinan terminus of arabinogalactan (AG) but are also present as extractable “free” lipids within the cell wall, mainly associated with trehalose to form trehalose dimycolate (TDM), also known as cord factor ( 3 ). Recent studies have also demonstrated the presence of free mycolates associated with M. tuberculosis biofilms ( 4 ). The crucial importance of the cell envelope integrity for the viability of M. tuberculosis has raised interest in understanding the enzymatic pathway for mycolic acid biosynthesis ( 5 ). Our knowledge of the biosynthesis of mycolic acid is important for finding new therapeutic targets to combat tuberculosis as well as for unraveling the mode of action of several existing antitubercular drugs ( 6 – 8 ). Indeed, the inhibition of mycolic acid biosynthesis is the primary effect of the frontline drug isoniazid (INH) ( 9 ). This unique metabolic pathway represents an important and attractive reservoir of targets for future chemotherapy, whose development is particularly urgent in the context of multidrug-resistant (MDR) tuberculosis and the nearly untreatable ( 10 ) extensively drug-resistant (XDR) strains of M. tuberculosis.
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Structures of representative mycolic acids in M. tuberculosis. General scheme of mycolic acid pyrolytic cleavage to form the α branch and meroaldehyde (top panel). Representative mycolic acid structures are named (left). Polymethylenic parts of the meromycolate are marked with thin arrows and letters: a, b, c. The chain modifications are shown and annotated with the methyltransferase responsible for their synthesis. Two enzymes are listed without parentheses when the modification is lost only in a double mutant of the genes encoding the listed enzymes and not in either of the single mutants. The parenthetical enzyme plays a secondary role that is evident only when the gene encoding the primary enzyme is deleted.
Genomic organization of fas II operons in M. tuberculosis. Genes encoding the FAS II components are shown in gray, whereas the transcriptional repressor MabR is represented in black.
The FAS I and FAS II pathways in M. tuberculosis. In both systems, the chain elongation steps consist of an iterative series of reactions built on successive addition of a two-carbon unit to a nascent acyl group, and reaction intermediates are covalently linked to the acyl carrier protein AcpM. FAS I is capable of de novo synthesis from acetyl-CoA producing acyl-CoA either used to synthesize the α-branch or C16/C18-CoA that are directly shuttled into FAS II for the production of the meromycolic acid. FAS II is primed by the CoA-dependent β-ketoacyl-AcpM synthase FabH, which condenses the acyl-CoA with malonyl-AcpM to generate a β-ketoacyl-AcpM, subsequently converted into a saturated enoyl-AcpM by the sequential actions of a β-ketoacyl-AcpM reductase (MabA), a β-hydroxyacyl-AcpM dehydratase complex (HadABC), and a trans-2-enoyl-AcpM reductase (InhA). Subsequent rounds of elongation are initiated by either the KasA or KasB β-ketoacyl AcpM synthases. KasA is thought to be responsible for the early rounds of elongations, whereas KasB is involved in later stages. Also shown are the acetyl-CoA carboxylase (AccA3/AccD6) that produces malonyl-CoA, the malonyl-CoA:AcpM transacylase (FabD) responsible for the synthesis of malonyl-AcpM, as well as the set of SAM-dependent methyltransferases involved in functionalization of the meromycolic acid.
Scheme of the FadD32-Pks13-AccD4 interplay during mycolic condensation. Prior to condensation the two acyl chains have to be activated (step 1). FadD32, a fatty acyl-AMP ligase (FAAL) converts the meromycolyl-AcpM to meromycolyl-AMP. AccD4 associating with the AccA3 carboxylates acyl-CoA yielding carboxyacyl-CoA. Both substrates are then loaded onto Pks13 (step 2). The meromycolyl-AMP is transacylated onto the Pks13 N-terminal ACP domain by FadD32 fatty acyl-ACP synthetase (FAAS) activity and subsequently transferred onto the ketoacyl synthase (KS) domain. The carboxyacyl-CoA initially binds to the Pks13 acyl transferase (AT) domain, which catalyzes its transfer onto the Pks13 C-terminal ACP. The KS domain catalyzes the Claisen-type condensation between the meromycolyl and the carboxyacyl chains to produce an α-alkyl β-keto thioester, which remains bound to the C-terminal ACP domain (step 3). The thioesterase (TE) catalyzes the release of the α-alkyl β-ketoacyl chain and its transfer onto an unknown acceptor (X1) (step 4). Finally, the reduction of the β-ketoacyl product by the CmrA reductase leads to the mature mycolic acid, which is transferred onto another unknown acceptor (X2) (step 5).
Genes involved in M. tuberculosis fatty/mycolic acid biosynthesis