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Systems metabolic engineering of Escherichia coli for gram scale production of the antitumor drug deoxyviolacein from glycerol
Author(s) -
Rodrigues André Luis,
Becker Judith,
de Souza Lima André Oliveira,
Porto Luismar Marques,
Wittmann Christoph
Publication year - 2014
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.25297
Subject(s) - escherichia coli , pentose , glycerol , biochemistry , pentose phosphate pathway , catabolite repression , arabinose , biosynthesis , biology , catabolism , metabolic engineering , fermentation , tryptophan , chemistry , microbiology and biotechnology , metabolism , enzyme , xylose , gene , amino acid , glycolysis , mutant
Deoxyviolacein is a microbial drug with biological activity against tumors, gram‐positive bacteria, and fungal plant pathogens. Here, we describe an Escherichia coli strain for heterologous production of this high‐value drug from glycerol. Plasmid‐based expression of the deoxyviolacein cluster vioABCE was controlled by the araBAD promoter and induction by L ‐arabinose. Through elimination of L ‐arabinose catabolism in E. coli , the pentose sugar could be fully directed to induction of deoxyviolacein biosynthesis and was no longer metabolized, as verified by 13 C isotope experiments. Deletion of the araBAD genes beneficially complemented with previously described (i) engineering of the pentose phosphate pathway, (ii) chorismate biosynthesis, (iii) tryptophan biosynthesis, (iv) improved supply of L ‐serine, (v) elimination of tryptophan repression, and (vi) of tryptophan catabolism. Subsequent screening of the created next‐generation producer E. coli dVio‐8 identified glycerol as optimum carbon source and a level of 100 mg L −1 of L ‐arabinose as optimum for induction. Transferred to a glycerol‐based fed‐batch process, E. coli dVio‐8 surpassed the gram scale and produced 1.6 g L −1 deoxyviolacein. With straightforward extraction from culture broth and purification by flash chromatography, deoxyviolacein was obtained at >99.5% purity. Biotechnol. Bioeng. 2014;111: 2280–2289. © 2014 Wiley Periodicals, Inc.