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Chem Biol. 2004 Nov;11(11):1573-82.

Enhanced macrocyclizing activity of the thioesterase from tyrocidine synthetase in presence of nonionic detergent.

Author information

1
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 240 Longwood Avenue, Boston, MA 02115, USA.

Abstract

Macrocyclization carried out by thioesterase domains of multimodular nonribosomal peptide synthetases (NRPSs) is a key step in the biosynthesis of many biologically active peptides. The thioesterase excised from tyrocidine synthetase is a versatile macrocyclization catalyst and a useful tool for chemoenzymatic synthesis of diverse cyclic peptides. However, its utility is limited by its short lifetime of catalytic activity as well as significant flux of the acyl-enzyme intermediate to hydrolysis. The addition of Brij 58, a nonionic detergent, above the critical micelle concentration, has dramatic effects on enzyme activity: catalytic activity is extended to >60 min and the rate of cyclization (but not hydrolysis) increases 6-fold, resulting in a net 150- to 300-fold increase in cyclic product yields. This enhanced activity allowed enzymatic macrocyclization of a solid phase library of tyrocidine decapeptides to identify acceptable substitutions at the Orn9 position which had previously been inaccessible for diversification.

PMID:
15556008
DOI:
10.1016/j.chembiol.2004.09.003
[Indexed for MEDLINE]
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