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• 作者：Hiroyuki Nozaki ; Shinji Kuroda ; Kunihiko Watanabe ; Kenzo Yokozeki
• 关键词：α ; -Methyl-l-serine ; α ; -Methylserine hydroxymethyltransferase ; Pyridoxal-5′ ; -phosphate ; Tetrahydrofolate
• 刊名：Journal of Molecular Catalysis B: Enzymatic
• 出版年：2009
• 出版时间：April 2009
• 年：2009
• 卷：56
• 期：4
• 页码：221-226
• 全文大小：391 K

文摘

Through the screening of microorganisms capable of utilizing α-methylserine, three representative strains belonging to the bacterial genera Paracoccus, Aminobacter, and Ensifer were selected as potent producers of α-methylserine hydroxymethyltransferase, an enzyme that catalyzes the interconversion between α-methyl-l-serine and d-alanine via tetrahydrofolate. Among these strains, Paracoccus sp. AJ110402 was selected as the strain exhibiting the highest α-methylserine hydroxymethyltransferase activity. The enzyme was purified to homogeneity from a cell-free extract of this strain. The native enzyme is a homodimer with apparent molecular mass of 85 kDa and contains 1 mol of pyridoxal-5′-phosphate per mol of the subunit. The K_m for α-methyl-l-serine and tetrahydrofolate was 0.54 mM and 73 μM, respectively. The gene from Paracoccus sp. AJ110402 encoding α-methylserine hydroxymethyltransferase was cloned and expressed in Escherichia coli. Sequence analysis revealed an open reading frame of 1278 bp, encoding a polypeptide with a calculated molecular mass of 46.0 kDa. Using E. coli cells as whole-cell catalysts, 9.7 mmol of α-methyl-l-serine was stereoselectively obtained from 15 mmol of d-alanine and 13.2 mmol of formaldehyde.