In Vitro Characterization of a Heterologously Expressed Nonribosomal Peptide Synthetase Involved in Phosphinothricin Tripeptide Biosynthesis

Overview

Journal Biochemistry

Specialty Biochemistry

Date 2009 May 13

PMID 19432442

Citations 9

Authors

Jin-Hee Lee

Bradley S Evans

Gongyong Li

Neil L Kelleher

Wilfred A van der Donk

Affiliations

Soon will be listed here.

Abstract

The late stages of biosynthesis of phosphinothricin tripeptide (PTT) involve peptide formation and methylation on phosphorus. The exact timing of these transformations is not known. To provide insight into this question, we developed a heterologous expression system for PhsA, one of three NRPS proteins in PTT biosynthesis. The apparent k(cat)/K(m) value for ATP-pyrophosphate exchange activity for d,l-N-acetylphosphinothricin was 3.5 muM(-1) min(-1), whereas the k(cat)/K(m,app) for l-N-acetyldemethylphosphinothricin was 0.5 microM(-1) min(-1), suggesting the former might be the physiological substrate. Each substrate could be loaded onto the phosphopantetheine arm of the thiolation domain as observed by Fourier transform mass spectrometry (FTMS).

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