Abstract
The C-1027 enediyne antibiotic contains an unusual 3-chloro-4,5-dihydroxy-beta-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. The sgcC4 gene encodes a protein with strong homology to dehydroalanine-dependent histidine/phenylalanine ammonia lyases. The sgcC4 gene was expressed in E. 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 of l-tyrosine to (S)-beta-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 in kcat/KM. SgcC4 represents a novel type of aminomutase, extending the known MIO chemistry from ammonia lyases into aminomutases.
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