Engineered urdamycin glycosyltransferases are broadened and altered in substrate specificity
Engineered urdamycin glycosyltransferases are broadened and altered in substrate specificity
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DOI：
10.1016/s1074-5521(02)00114-x
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发表时间：
2002-03-01
影响因子：
--
通讯作者：
Bechthold, A
中科院分区：
文献类型：
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作者：
Hoffmeister, D;Wilkinson, B;Bechthold, A
组合生物合成是一种很有前途的技术，可以为药物开发提供修饰的天然产物。糖基转移酶是在糖苷元生物合成和糖衍生化之间用酶法连接的工具。为了克服它们固有的特异性所设置的局限性，我们对两个乌尔达霉素脱氧糖糖基转移酶UrdGT1b和UrdGT1c的核苷酸糖和受体底物特异性的蛋白质区域进行了基因工程改造。靶向氨基酸交换将潜在决定底物特异性的氨基酸数量减少到10个。随后，创建了一个基因库，使得来自两个亲本基因的这十个氨基酸的密码子只被独立地组合在一起。文库成员表现出亲本和/或新的特异性，后者负责乌尔达霉素P的生物合成，该侧链带有迄今未知的乌尔达霉素。
Combinatorial biosynthesis is a promising technique used to provide modified natural products for drug development. To enzymatically bridge the gap between what is possible in aglycon biosynthesis and sugar derivatization, glycosyltransferases are the tools of choice. To overcome limitations set by their intrinsic specificities, we have genetically engineered the protein regions governing nucleotide sugar and acceptor substrate specificities of two urdamycin deoxysugar glycosyltransferases, UrdGT1b and UrdGT1c. Targeted amino acid exchanges reduced the number of amino acids potentially dictating substrate specificity to ten. Subsequently, a gene library was created such that only codons of these ten amino acids from both parental genes were independently combined. Library members displayed parental and/or a novel specificity, with the latter being responsible for the biosynthesis of urdamycin P that carries a branched saccharide side chain hitherto unknown for urdamycins.