Chemoenzymatic approach to enantiopure streptogramin B variants: Characterization of stereoselective pristinamycin I cyclase from Streptomyces pristinaespiralis

Christoph Mahlert, Stephan A. Sieber, Jan Grünewald, Mohamed A. Marahiel

Research output: Contribution to journalArticlepeer-review

32 Scopus citations

Abstract

Streptogramin B antibiotics are cyclic peptide natural products produced by Streptomyces species. In combination with the synergistic group A component, they are "last line of defense" antimicrobial agents against multiresistant cocci. The racemization sensitivity of the phenylglycine (Phg7) ester is a complex challenge in total chemical synthesis of streptogramin B molecules. To provide fast and easy access to novel streptogramin antibiotics, we introduce a novel chemoenzymatic strategy in which diversity is generated by standard solid phase protocols and stereoselectivity by subsequent enzymatic cyclization. For this approach, we cloned, overproduced, and biochemically characterized the recombinant thioesterase domain SnbDE TE of the pristinamycin I nonribosomal peptide synthetase from Streptomyces pristinaespiralis. SnbDE TE catalyzes regioselective ring closure of linear peptide thioester analogues of pristinamycin I as well as stereoselective cyclization out of complex in situ racemizing substrate mixtures, enabling synthesis of Streptogramin B variants via a dynamic kinetic resolution assay. A remarkable substrate tolerance was detected for the enzymatic cyclization including all the seven positions of the peptide backbone. Interestingly, SnbDE TE was observed to be the first cyclase from a macrolactone forming NRPS which is additionally able to catalyze macrolactamization of peptide thioester substrates. An N-methylated peptide bond between positions 4 and 5 is mandatory for a high substrate turnover. The presented strategy is potent to screen for analogues with improved activity and guides our understanding of structure-activity relationships in the important class of streptogramin antibiotics.

Original languageEnglish
Pages (from-to)9571-9580
Number of pages10
JournalJournal of the American Chemical Society
Volume127
Issue number26
DOIs
StatePublished - 6 Jul 2005
Externally publishedYes

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