317-66-8Relevant articles and documents
Investigating the molecular determinants for substrate channeling in BphI-BphJ, an aldolase-dehydrogenase complex from the polychlorinated biphenyls degradation pathway
Carere, Jason,Baker, Perrin,Seah, Stephen Y. K.
, p. 8407 - 8416 (2011)
BphI-BphJ, an aldolase-dehydrogenase complex from the polychlorinated biphenyls (PCBs) degradation pathway, cleaves 4-hydroxy-2-oxoacids to pyruvate and an aldehyde. The enzyme complex was shown to exhibit substrate channeling, whereby linear aldehydes of up to 6 carbons long and branched isobutyraldehyde were directly channeled from the aldolase to the dehydrogenase with greater than 80% efficiency. BphI variants G322F, G322L, and G323F were created and were found to block aldehyde channeling. The dehydrogenase cofactor NADH was able to activate the catalytic activity of the aldol cleavage reaction in these variants, suggesting that activation of BphI by BphJ cofactors is not solely due to faster aldehyde release. A G323L variant was able to channel acetaldehyde but not the larger propionaldehyde while the G323A variant was able to channel butyraldehyde but not its isomer isobutyraldehyde, confirming that the restricted channeling of aldehydes in these glycine variants are due to steric blockage of the channel. Substitution of His-20 and Tyr-290 in BphI led to significant reductions in aldehyde channeling efficiencies. A mechanism of substrate channeling involving these two gating residues is proposed.
Crystal structures of Acetobacter aceti succinyl-coenzyme A (CoA):Acetate CoA-transferase reveal specificity determinants and illustrate the mechanism used by class i CoA-transferases
Mullins, Elwood A.,Kappock, T. Joseph
, p. 8422 - 8434 (2013/01/15)
Coenzyme A (CoA)-transferases catalyze transthioesterification reactions involving acyl-CoA substrates, using an active-site carboxylate to form covalent acyl anhydride and CoA thioester adducts. Mechanistic studies of class I CoA-transferases suggested that acyl-CoA binding energy is used to accelerate rate-limiting acyl transfers by compressing the substrate thioester tightly against the catalytic glutamate [White, H., and Jencks, W. P. (1976) J. Biol. Chem. 251, 1688-1699]. The class I CoA-transferase succinyl-CoA:acetate CoA-transferase is an acetic acid resistance factor (AarC) with a role in a variant citric acid cycle in Acetobacter aceti. In an effort to identify residues involved in substrate recognition, X-ray crystal structures of a C-terminally His6-tagged form (AarCH6) were determined for several wild-type and mutant complexes, including freeze-trapped acetylglutamyl anhydride and glutamyl-CoA thioester adducts. The latter shows the acetate product bound to an auxiliary site that is required for efficient carboxylate substrate recognition. A mutant in which the catalytic glutamate was changed to an alanine crystallized in a closed complex containing dethiaacetyl-CoA, which adopts an unusual curled conformation. A model of the acetyl-CoA Michaelis complex demonstrates the compression anticipated four decades ago by Jencks and reveals that the nucleophilic glutamate is held at a near-ideal angle for attack as the thioester oxygen is forced into an oxyanion hole composed of Gly388 NH and CoA N2″. CoA is nearly immobile along its entire length during all stages of the enzyme reaction. Spatial and sequence conservation of key residues indicates that this mechanism is general among class I CoA-transferases.
Establishing a toolkit for precursor-directed polyketide biosynthesis: Exploring substrate promiscuities of acid-CoA ligases
Go, Maybelle Kho,Chow, Jeng Yeong,Cheung, Vivian Wing Ngar,Lim, Yan Ping,Yew, Wen Shan
experimental part, p. 4568 - 4579 (2012/08/28)
Polyketides are chemically diverse and medicinally important biochemicals that are biosynthesized from acyl-CoA precursors by polyketide synthases. One of the limitations to combinatorial biosynthesis of polyketides has been the lack of a toolkit that describes the means of delivering novel acyl-CoA precursors necessary for polyketide biosynthesis. Using five acid-CoA ligases obtained from various plants and microorganisms, we biosynthesized an initial library of 79 acyl-CoA thioesters by screening each of the acid-CoA ligases against a library of 123 carboxylic acids. The library of acyl-CoA thioesters includes derivatives of cinnamyl-CoA, 3-phenylpropanoyl-CoA, benzoyl-CoA, phenylacetyl-CoA, malonyl-CoA, saturated and unsaturated aliphatic CoA thioesters, and bicyclic aromatic CoA thioesters. In our search for the biosynthetic routes of novel acyl-CoA precursors, we discovered two previously unreported malonyl-CoA derivatives (3-thiophenemalonyl-CoA and phenylmalonyl-CoA) that cannot be produced by canonical malonyl-CoA synthetases. This report highlights the utility and importance of determining substrate promiscuities beyond conventional substrate pools and describes novel enzymatic routes for the establishment of precursor-directed combinatorial polyketide biosynthesis. (Chemical Presented).
