Optimizing dimodular nonribosomal peptide synthetases and natural dipeptides in an Escherichia coli heterologous host
Nonribosomal peptides are an important class of natural products that have a broad range of biological activities. Their structural complexity often prevents simple chemical synthesis, and production from the natural producer is often low, which deters pharmaceutical development. Expression
of biosynthetic machinery in heterologous host organisms like Escherichia coli is one way to access these structures, and subsequent optimization of these systems is critical for future development. We utilized the aureusimine biosynthetic gene cluster as a model system to identify
the optimal conditions to produce nonribosomal peptides in the isopropyl β-d-1-thiogalactopyranoside (IPTG)-inducible T7 promoter system of pET28. Single reaction monitoring of nonribosomal products was used to find the optimal concentration of IPTG, postinduction temperature, and the
effect of amino acid precursor supplementation. In addition, principle component analysis of these extracts identified 3 previously undiscovered pyrazine products of the aureusimine biosynthetic locus, highlighting the utility of heterologously expressing nonribosomal peptide synthetases to
find new products.
Keywords: aureusimine; biosynthesis; biosynthèse; expression hétérologue; heterologous expression; nonribosomal peptide; peptide non ribosomaux; pyrazine
Document Type: Research Article
Affiliations: Department of Biochemistry and Biomedical Sciences, McMaster University, 1280 Main St. W, Hamilton, ON, L8N 3Z5, Canada.
Publication date: 01 January 2013
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