Catabolic alanine racemase from Salmonella typhimurium: DNA sequence, enzyme purification, and characterization.
Catabolic alanine racemase from Salmonella typhimurium: DNA sequence, enzyme purification, and characterization.
复制标题
DOI：
10.1021/bi00317a015
复制
发表时间：
1984-10
期刊：
影响因子：
2.9
通讯作者：
S. Wasserman;E. Daub;P. Grisafi;D. Botstein;C. Walsh
中科院分区：
文献类型：
--
作者：
S. Wasserman;E. Daub;P. Grisafi;D. Botstein;C. Walsh
将鼠伤寒沙门氏菌dadB基因编码的丙氨酸外消旋酶从一株高产菌株中纯化到90%的同源性。在37℃时，该酶的比活为1400U/mg(Vmax，L-到D-丙氨酸)。活性酶分子是分子量为39000的单体，每个亚基结合一个吡哆醛5‘-磷酸分子。L-丙氨酸和D-丙氨酸的Km分别为8.2mM和2.1mM。对成交额的衡量得出了预期的Keq值为1.0。对纯化多肽的25个N端氨基酸残基中的22个氨基酸残基的测定使编码该结构基因的克隆DNA得以定位。亚克隆DNA测序表明，该基因编码356个氨基酸的多肽，其计算相对分子质量(脱辅酶)为39044。
The alanine racemase encoded by the Salmonella typhimurium dadB gene was purified to 90% homogeneity from an overproducing strain. At 37 degrees C the enzyme has a specific activity of 1400 units/mg (V max, L- to D-alanine). Active enzyme molecules are monomers of Mr 39 000 with one molecule of pyridoxal 5'-phosphate bound per subunit. The Km's for L- and D-alanine are 8.2 and 2.1 mM, respectively. Measurement of turnover numbers yielded the expected Keq value of 1.0. Determination of 22 of the 25 N-terminal amino acid residues of the purified polypeptide allowed localization of cloned DNA encoding the structural gene. Sequencing of subcloned DNA revealed that the dadB gene encodes a polypeptide of 356 amino acids whose calculated molecular weight (apoenzyme) was 39 044.