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Expanding metabolism for biosynthesis of nonnatural alcohols.

ABSTRACT: Nature uses a limited set of metabolites to perform all of the biochemical reactions. To increase the metabolic capabilities of biological systems, we have expanded the natural metabolic network, using a nonnatural metabolic engineering approach. The branched-chain amino acid pathways are extended to produce abiotic longer chain keto acids and alcohols by engineering the chain elongation activity of 2-isopropylmalate synthase and altering the substrate specificity of downstream enzymes through rational protein design. When introduced into Escherichia coli, this nonnatural biosynthetic pathway produces various long-chain alcohols with carbon number ranging from 5 to 8. In particular, we demonstrate the feasibility of this approach by optimizing the biosynthesis of the 6-carbon alcohol, (S)-3-methyl-1-pentanol. This work demonstrates an approach to build artificial metabolism beyond the natural metabolic network. Nonnatural metabolites such as long chain alcohols are now included in the metabolite family of living systems.

SUBMITTER: Zhang K

PROVIDER: S-EPMC2634914 | biostudies-literature | 2008 Dec

REPOSITORIES: biostudies-literature

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Expanding metabolism for biosynthesis of nonnatural alcohols.

Zhang Kechun K Sawaya Michael R MR Eisenberg David S DS Liao James C JC

Proceedings of the National Academy of Sciences of the United States of America 20081208 52

Nature uses a limited set of metabolites to perform all of the biochemical reactions. To increase the metabolic capabilities of biological systems, we have expanded the natural metabolic network, using a nonnatural metabolic engineering approach. The branched-chain amino acid pathways are extended to produce abiotic longer chain keto acids and alcohols by engineering the chain elongation activity of 2-isopropylmalate synthase and altering the substrate specificity of downstream enzymes through r ...[more]

PMID: 19064911

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Project description:The expansion of the genetic repertoire of an organism by gene duplication or horizontal gene transfer (HGT) can aid adaptation. Streptomyces bacteria are prolific producers of bioactive specialized metabolites that have adaptive functions in nature and have found extensive utility in human medicine. While the biosynthesis of these specialized metabolites is directed by dedicated biosynthetic gene clusters, little attention has been focused on how these organisms have evolved robustness in their genomes to facilitate the metabolic plasticity required to provide chemical precursors for biosynthesis during the complex metabolic transitions from vegetative growth to specialized metabolite production and sporulation. Here, we examine genetic redundancy in actinobacteria and show that specialized metabolite-producing bacterial families exhibit gene family expansion in primary metabolism. Focusing on a gene duplication event, we show that the two pyruvate kinases in the genome of Streptomyces coelicolor arose by an ancient duplication event and that each has evolved altered enzymatic kinetics, with Pyk1 having a 20-fold-higher kcat than Pyk2 (4,703 s-1 compared to 215 s-1, respectively), and yet both are constitutively expressed. The pyruvate kinase mutants were also found to be compromised in terms of fitness compared to wild-type Streptomyces These data suggest that expanding gene families can help maintain cell functionality during metabolic perturbation such as nutrient limitation and/or specialized metabolite production.IMPORTANCE The rise of antimicrobial-resistant infections has prompted a resurgence in interest in understanding the production of specialized metabolites, such as antibiotics, by Streptomyces The presence of multiple genes encoding the same enzymatic function is an aspect of Streptomyces biology that has received little attention; however, understanding how the metabolic expansion influences these organisms can help enhance production of clinically useful molecules. Here, we show that expanding the number of pyruvate kinases enables metabolic adaptation, increases strain fitness, and represents an excellent target for metabolic engineering of industrial specialized metabolite-producing bacteria and the activation of cryptic specialized metabolites.

Dataset Information

Expanding metabolism for biosynthesis of nonnatural alcohols.

Publications

Expanding metabolism for biosynthesis of nonnatural alcohols.

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