title: Analysis of a parallel branch in the mitomycin biosynthetic pathway involving the mitN-encoded aziridine N-methyltransferase.
abstract: Mitomycin C is a natural product with potent alkylating activity, and it is an important anticancer drug and antibiotic. mitN, one of three genes with high similarity to methyltransferases, is located within the mitomycin biosynthetic gene cluster. An inframe deletion in mitN of the mitomycin biosynthetic pathway was generated in Streptomyces lavendulae to produce the DHS5373 mutant strain. Investigation of DHS5373 revealed continued production of mitomycin A and mitomycin C in addition to the accumulation of a new mitomycin analog, 9-epi-mitomycin C. The mitN gene was overexpressed in Escherichia coli, and the histidine-tagged protein (MitN) was purified to homogeneity. Reaction of 9-epi-mitomycin C with MitN in the presence of S-adenosylmethionine yielded mitomycin E showing that the enzyme functions as an aziridine N-methyltransferase. Likewise, MitN was also shown to convert mitomycin A to mitomycin F under the same reaction conditions. We conclude that MitN plays an important role in a parallel biosynthetic pathway leading to the subclass of mitomycins with 9alpha-stereochemistry but is not involved directly in the biosynthesis of mitomycins A and C.
authors.affiliation: Life Sciences Institute and Department of Medicinal Chemistry, University of Michigan, Ann Arbor, Michigan 48109, USA.
authors.forename: Namthip
authors.initials: N
authors.lastname: Sitachitta
authors.forename: Nicole B
authors.initials: NB
authors.lastname: Lopanik
authors.forename: Yingqing
authors.initials: Y
authors.lastname: Mao
authors.forename: David H
authors.initials: DH
authors.lastname: Sherman
mesh_terms: D001388:Aziridines / Q000737:chemistry*; D002412:Cations; D003001:Cloning, Molecular; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000378:metabolism; D017353:Gene Deletion; D006639:Histidine / Q000737:chemistry; D007700:Kinetics; D008780:Methyltransferases / Q000096:biosynthesis / Q000737:chemistry*; D016685:Mitomycin / Q000096:biosynthesis* / Q000737:chemistry; D008937:Mitomycins / Q000737:chemistry; D008956:Models, Chemical; D008968:Molecular Conformation; D013302:Streptomyces / Q000378:metabolism*
chemical_list: D001388:Aziridines; D002412:Cations; D008937:Mitomycins; D006639:Histidine; D016685:Mitomycin; C033132:aziridine; C046000:mitomycin A; D008780:Methyltransferases; C522739:aziridine N-methyltransferase, Streptomyces lavendulae