Graduate School of Pharmaceutical Sciences, Kitasato University, 5-9-1 Shirokane, Minato-ku, Tokyo, 108-8641, Japan.
Amino Acids. 2017 Nov;49(11):1885-1894. doi: 10.1007/s00726-017-2486-2. Epub 2017 Sep 11.
The peptidoglycan layer of the bacterial cell wall typically contains D-alanine (D-Ala) and D-glutamic acid (D-Glu), and also various non-canonical D-amino acids that have been linked to peptidoglycan remodeling, inhibition of biofilm formation, and triggering of biofilm disassembly. Bacteria produce D-amino acids when adapting to environmental changes as a common survival strategy. In our previous study, we detected non-canonical D-amino acids in Escherichia coli grown in minimal medium. However, the biosynthetic pathways of non-canonical D-amino acids remain poorly understood. In the present study, we identified amino acid racemases in E. coli MG1655 (YgeA) and Bacillus subtilis (RacX) that produce non-canonical D-amino acids other than D-Ala and D-Glu. We characterized their enzymatic properties, and both displayed broad substrate specificity but low catalytic activity. YgeA preferentially catalyzes the racemization of homoserine, while RacX preferentially racemizes arginine, lysine, and ornithine. RacX is dimeric, and appears not to require pyridoxal 5'-phosphate (PLP) as a coenzyme as is the case with YgeA. To our knowledge, this is the first report on PLP-independent amino acid racemases possessing broad substrate specificity in E. coli and B. subtilis.
细菌细胞壁的肽聚糖层通常含有 D-丙氨酸 (D-Ala) 和 D-谷氨酸 (D-Glu)，以及各种与肽聚糖重塑、生物膜形成抑制和生物膜解体触发相关的非典型 D-氨基酸。细菌在适应环境变化时会产生 D-氨基酸，这是一种常见的生存策略。在我们之前的研究中，我们在最小培养基中生长的大肠杆菌中检测到非典型 D-氨基酸。然而，非典型 D-氨基酸的生物合成途径仍知之甚少。在本研究中，我们在大肠杆菌 MG1655（YgeA）和枯草芽孢杆菌（RacX）中鉴定出了产生非典型 D-氨基酸（除了 D-Ala 和 D-Glu）的氨基酸消旋酶。我们对它们的酶学性质进行了表征，两者都表现出广泛的底物特异性，但催化活性较低。YgeA 优先催化高丝氨酸的消旋化，而 RacX 优先催化精氨酸、赖氨酸和鸟氨酸的消旋化。RacX 是二聚体，似乎不需要吡哆醛 5'-磷酸 (PLP) 作为辅酶，而 YgeA 需要 PLP 作为辅酶。据我们所知，这是首次在大肠杆菌和枯草芽孢杆菌中报道具有广泛底物特异性的非依赖 PLP 的氨基酸消旋酶。
