A novel 4-methylideneimidazole-5-one-containing tyrosine aminomutase in enediyne antitumor antibiotic C-1027 biosynthesis
A novel 4-methylideneimidazole-5-one-containing tyrosine aminomutase in enediyne antitumor antibiotic C-1027 biosynthesis
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DOI：
10.1021/ja034609m
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发表时间：
2003-05-21
影响因子：
15
通讯作者：
Shen, B
中科院分区：
文献类型：
--
作者：
Christenson, SD;Liu, W;Shen, B
C-1027烯二炔抗生素含有一个不寻常的3-氯-4，5-二羟基-β-苯丙氨酸部分，被认为是通过氨基变位酶反应从酪氨酸衍生而来。然而，在C-1027基因簇内鉴定的基因都不编码与已知氨基变位酶具有强同源性的蛋白质。ThesgcC 4基因编码的蛋白与脱氢丙氨酸依赖性组氨酸/苯丙氨酸氨裂解酶具有很强的同源性。sgcC 4基因在E.将过量产生的SgcC 4纯化为His 6标记的融合蛋白。纯化的SgcC 4的生化特征确定SgcC 4是一种氨基变位酶，催化l-酪氨酸转化为（S）-β-酪氨酸，并在其活性位点使用4-亚甲基咪唑-5-酮（MIO）。后者得到了硼氢化物和氰化物抑制研究的支持，并通过定点诱变证实。S153 A突变体表现出340倍的降低inkcat/KM。SgcC 4代表一种新型的氨基变位酶，将已知的MIO化学从氨裂解酶扩展到氨基变位酶。
The C-1027 enediyne antibiotic contains an unusual 3-chloro-4,5-dihydroxy-β-phenylalanine moiety that is thought to be derived from tyrosine by an aminomutase reaction. However, none of the genes identified within the C-1027 gene cluster encode proteins with strong homology to known aminomutases. ThesgcC4gene encodes a protein with strong homology to dehydroalanine-dependent histidine/phenylalanine ammonia lyases. The sgcC4 gene was expressed inE. coli, and overproduced SgcC4 was purified as a His6-tagged fusion protein. Biochemical characterization of the purified SgcC4 establishes that SgcC4 is an aminomutase that catalyzes the conversion ofl-tyrosine to (S)-β-tyrosine and employs 4-methylideneimidazole-5-one (MIO) at its active site. The latter was supported by borohydride and cyanide inhibition studies and confirmed by site-directed mutagenesis. The S153A mutant exhibited a 340-fold decrease inkcat/KM. SgcC4 represents a novel type of aminomutase, extending the known MIO chemistry from ammonia lyases into aminomutases.