The enediyne antibiotics are the focus of intense research activity in the fields of chemistry, biology, and medical sciences because of their unique molecular architectures, biological activities, and modes of action (, ). Since the unveiling of the structure of neocarzinostatin chromophore () in 1985, the enediyne family has grown steadily. Thus far, there have been three basic groups within the enediyne antibiotic family: (i) the calicheamicin-esperamicin type, which includes the calicheamicins, the esperamicins, and namenamicin; (ii) the dynemicin type; and (iii) the chromoprotein type, consisting of an apoprotein and an unstable enediyne chromophore. The last group includes neocarzinostatin, kedarcidin, C-1027 (Fig. ), and maduropeptin, whose enediyne chromophore structures have been established, as well as several others whose enediyne chromophore structures are yet to be determined due to their instabilities (). N1999A2, in contrast to the other chromoproteins, exists as an enediyne chromophore alone, even though its structure is very similar to those of the other chromoprotein chromophores ().
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. 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)-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 in kcat/KM. SgcC4 represents a novel type of aminomutase, extending the known MIO chemistry from ammonia lyases into aminomutases.
Title: Biosynthesis of Enediyne Antitumor Antibiotics
32. Discovery of a free-standing condensation enzyme that can catalyze both C-O ester and C-N amide bond formation for the biosynthesis of the C-1027 enediyne antitumor antibiotic (2009, 4183-4188)
Biosynthesis of the enediyne antitumor antibiotic C-1027 ..
C-1027, the most potent member of the enediyne antitumor antibiotic family, is produced by Streptomyces globisporus C-1027 and consists of an apoprotein (encoded by the cagA gene) and a nonpeptidic chromophore. The C-1027 chromophore could be viewed as being derived biosynthetically from a benzoxazolinate, a deoxyamino hexose, a β-amino acid, and an enediyne core. By adopting a strategy for cloning of the C-1027 biosynthesis gene cluster by mapping a putative dNDP-glucose 4,6-dehydratase (NGDH) gene to cagA, we have localized 75 kb of contiguous DNA from S. globisporus. DNA sequence analysis of two regions of the cloned gene cluster revealed two genes, sgcA and sgcB, that encode an NGDH enzyme and a transmembrane efflux protein, respectively, and confirmed that the cagA gene resides approximately 14 kb upstream of the sgcAB locus. The involvement of the cloned gene cluster in C-1027 biosynthesis was demonstrated by disrupting the sgcA gene to generate C-1027-nonproducing mutants and by complementing the sgcA mutants in vivo to restore C-1027 production. These results represent the first cloning of a gene cluster for enediyne antitumor antibiotic biosynthesis and provide a starting point for future genetic and biochemical investigations of C-1027 biosynthesis.
(2002) Biosynthesis of the enediyne antitumor antibiotic C-1027
Horsman, Yihua Chen, Wenli Li, and Ben Shen (2009) Characterization of the SgcF epoxide hydrolase supporting a (R)-vicinal diol intermediate for enediyne antitumor antibiotic C-1027 biosynthesis.