The Mycobacterium tuberculosis Very-Long-Chain Fatty Acyl-CoA Synthetase: Structural Basis for Housing Lipid Substrates Longer than the Enzyme

Abstract

The Mycobacterium tuberculosis acid-induced operon MymA encodes the fatty acyl-CoA synthetase FadD13 and is essential for virulence and intracellular growth of the pathogen. Fatty acyl-CoA synthetases activate lipids before entering into the metabolic pathways and are also involved in transmembrane lipid transport. Unlike soluble fatty acyl-CoA synthetases, but like the mammalian integral-membrane very-long-chain acyl-CoA synthetases, FadD13 accepts lipid substrates up to the maximum length tested (C(26)). Here, we show that FadD13 is a peripheral membrane protein. The structure and mutational studies reveal an arginine- and aromatic-rich surface patch as the site for membrane interaction. The protein accommodates a hydrophobic tunnel that extends from the active site toward the positive patch and is sealed by an arginine-rich lid-loop at the protein surface. Based on this and previous data, we propose a structural basis for accommodation of lipid substrates longer than the enzyme and transmembrane lipid transport by vectorial CoA-esterification.

The Non-catalytic “Cap Domain” of a Mycobacterial Metallophosphoesterase Regulates Its Expression and Localization in the Cell

“…Therefore, the exact mechanism by which Rv0805 is exported remains unknown. Examples of other mycobacterial proteins that localize in a secretion-system independent manner to the cell envelope include FadD13 (26) and Rv1818c (35). In the case of Rv1818c, the first 30 amino acids of its PE (proline glutamate-rich) domain were both necessary and sufficient for localization to the cell envelope (35).…”

“…In the case of Rv1818c, the first 30 amino acids of its PE (proline glutamate-rich) domain were both necessary and sufficient for localization to the cell envelope (35). In the case of FadD13, however, docking to the cell membrane was determined by the distribution of positively charged and hydrophobic residues on the protein rather than a specific targeting domain (26).…”

Structures of Mycobacterium tuberculosis FadD10 Protein Reveal a New Type of Adenylate-forming Enzyme

“…3B, panel i) notably varies among adenylate-forming proteins. A short helix equivalent to A11 is found in only four members, including the adenylation domains DhbE (30) and DltA (35), as well as the acyl-CoA synthetases, long chain fatty acyl-CoA synthetase (32), and M. tuberculosis FACL13 (29). However, the glutamate residue (Glu-340) essential to forming inter-domain hydrogen bonds in M. tuberculosis FadD10 is replaced by nonpolar residues in these four homologous proteins (Fig.…”

“…The bottom of this groove is about 9.4 Å from C12 of the dodecanoyl chain, raising the possibility that the enzyme could accommodate a longer fatty acid substrate. We compared the fatty acid binding tunnels of the very long chain fatty acyl-CoA synthetase M. tuberculosis FACL13 (active on fatty acid with up to 26 carbons) (29), FadD10, and the human medium chain fatty acyl-CoA synthetase (active on fatty acid with up to 10 carbons) (33). The linear distances between the distal end and the portal of their fatty acyl binding tunnels are ϳ17, 14, and 10 Å, respectively, which indicates that the biological substrate of FadD10 is probably a long chain fatty acid.…”

Proteome-wide Subcellular Topologies of E. coli Polypeptides Database (STEPdb)

“…Additionally, we identified potential PIM proteins by their similarity to well characterized peripheral membrane proteins. We used FadD13 protein of Mycobacterium, that is an ATP-binding protein membrane associated through distinctive regions rich in aromatics (40). Six proteins have been identified as "By similarity" PIM proteins based on homology to FadD13 (EntE, Acs, CaiC, MenE, FadK, and PrpE).…”