1763-10-6Relevant articles and documents
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Vignais,Zabin
, p. 263,265 (1958)
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Substrate Recognition and Catalytic Mechanism of the Phosphate Acyltransferase PlsX from Bacillus subtilis
Jiang, Yiping,Qin, Mingming,Guo, Zhihong
, p. 2019 - 2028 (2020)
Phosphate: acyl-acyl carrier protein (ACP) acyltransferase PlsX is a peripheral enzyme catalysing acyl transfer to orthophosphate in phospholipid synthesis. Little is known about how it recognises substrates and catalyses the acyl transfer. Here we show that its active site includes many residues lining a long, narrow gorge at the dimeric interface, two positive residues forming a positive ACP docking pad next to the interfacial gorge, and a number of strictly conserved residues significantly contributing to the catalytic activity. These findings suggest a substrate recognition mode and a catalytic mechanism that are different from those of phosphotransacetylases catalysing a similar acyl transfer reaction. The catalytic mechanism involves substrate activation and transition-state stabilization by two strictly conserved residues, Lys184 and Asn229. Another noticeable feature of the catalysis is the release of the acyl phosphate product near the membrane, which might facilitate its membrane insertion.
Effect of carbon chain length in acyl coenzyme A on the efficiency of enzymatic transformation of okadaic acid to 7-O-acyl okadaic acid
Furumochi, Sachie,Onoda, Tatsuya,Cho, Yuko,Fuwa, Haruhiko,Sasaki, Makoto,Yotsu-Yamashita, Mari,Konoki, Keiichi
supporting information, p. 2992 - 2996 (2016/06/13)
Okadaic acid (OA), a product of dinoflagellate Prorocentrum spp., is transformed into 7-O-acyl OA in various bivalve species. The structural transformation proceeds enzymatically in vitro in the presence of the microsomal fraction from the digestive gland of bivalves. We have been using LC-MS/MS to identify OA-transforming enzymes by detecting 7-O-acyl OA, also known as dinophysistoxin 3 (DTX3). However, an alternative assay for DTX3 is required because the OA-transforming enzyme is a membrane protein, and surfactants for solubilizing membrane proteins decrease the sensitivity of LC-MS/MS. The present study examined saturated fatty acyl CoAs with a carbon chain length of 10 (decanoyl), 12 (dodecanoyl), 14 (tetradecanoyl), 16 (hexadecanoyl) and 18 (octadecanoyl) as the substrate for the in vitro acylation reaction. Saturated fatty acyl CoAs with a carbon chain length of 14, 16 and 18 exhibited higher yields than those with a carbon chain length of 10 or 12. Acyl CoAs with carbon chain lengths from 14 to 18 and containing either a diene unit, an alkyne unit, or an azide unit in the carbon chain were synthesized and shown to provide the corresponding DTX3 with a yield comparable to that of hexadecanoyl CoA. The three functional units can be conjugated with fluorescent reagents and are applicable to the development of a novel assay for DTX3.
Comparison of acyl-CoA synthetic activities and enantioselectivity toward 2-arylpropanoic acids in firefly luciferases
Kato, Dai-Ichiro,Yokoyama, Keisuke,Hiraishi, Yoshihiro,Takeo, Masahiro,Negoro, Seiji
experimental part, p. 1758 - 1762 (2012/02/02)
Measurement of thioesterification activities for dodecanoic acid (C12) and ketoprofen was done using five firefly luciferases, from Pyrocoelia miyako (PmL), Photinus pyralis (PpL), Luciola cruciata (LcL), Hotaria parvura (HpL), and Luciola mingrelica (LmL). Among these, PmL, PpL, and LcL showed the expected thioesterification activities toward both substrates. All the enzymes exhibited (R)-enantioselectivity toward ketoprofen, which had same tendency as firefly luciferase from Luciola lateralis (LUC-H). HpL and LmL, however, did not accept ketoprofen, although they had thioesterification activity toward C12. These results indicate that the substrate acceptance of luciferases for the thioesterification reaction varies dramatically relying on the origin of firefly. Hence we focused primarily on PmL and investigated the effect of pH on enzymatic activity. In addition, by determining the kinetic parameters at various pH values, we verified that the kcat parameter contributed to the preferential enantioselectivity of this enzyme.
