Modular enzyme assembly for enhanced cascade biocatalysis and metabolic flux

• Published in: Nature communications Vol. 10; no. 1; pp. 4248 - 11
• Main Authors: Kang, Wei, Ma, Tian, Liu, Min, Qu, Jiale, Liu, Zhenjun, Zhang, Huawei, Shi, Bin, Fu, Shuai, Ma, Juncai, Lai, Louis Tung Faat, He, Sicong, Qu, Jianan, Wing-Ngor Au, Shannon, Ho Kang, Byung, Yu Lau, Wilson Chun, Deng, Zixin, Xia, Jiang, Liu, Tiangang
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 18.09.2019; Nature Publishing Group; Nature Portfolio
• Subjects: 14/28; 38/1; 38/35; 38/39; 631/61/252; 631/61/318; 631/92/552; 82/58; 82/80; 82/83; Assembling; Biocatalysis; Bioreactors - microbiology; Biosynthetic Pathways - genetics; Carotenoids; Carotenoids - metabolism; Cascade chemical reactions; Catalysis; E coli; Enzymes; Escherichia coli - enzymology; Escherichia coli - genetics; Escherichia coli - metabolism; Humanities and Social Sciences; Industrial engineering; Lycopene; Lycopene - metabolism; Manufacturing engineering; Menaquinones; Metabolic Engineering - methods; Metabolic flux; Metabolism; Metabolites; Mevalonate pathway; Mevalonic acid; Mevalonic Acid - metabolism; multidisciplinary; Multienzyme complexes; Nanoparticles; Protein Engineering - methods; Saccharomyces cerevisiae; Saccharomyces cerevisiae - enzymology; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae - metabolism; Science; Science (multidisciplinary); Stoichiometry; Vitamin K 2 - metabolism
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-019-12247-w

Enzymatic reactions in living cells are highly dynamic but simultaneously tightly regulated. Enzyme engineers seek to construct multienzyme complexes to prevent intermediate diffusion, to improve product yield, and to control the flux of metabolites. Here we choose a pair of short peptide tags (RIAD and RIDD) to create scaffold-free enzyme assemblies to achieve these goals. In vitro, assembling enzymes in the menaquinone biosynthetic pathway through RIAD–RIDD interaction yields protein nanoparticles with varying stoichiometries, sizes, geometries, and catalytic efficiency. In Escherichia coli , assembling the last enzyme of the upstream mevalonate pathway with the first enzyme of the downstream carotenoid pathway leads to the formation of a pathway node, which increases carotenoid production by 5.7 folds. The same strategy results in a 58% increase in lycopene production in engineered Saccharomyces cerevisiae . This work presents a simple strategy to impose metabolic control in biosynthetic microbe factories. Metabolic enzymes often form supramolecular complexes to improve product yield. Here the authors use short peptide tags to create scaffold-free assemblies and synthetic metabolic nodes.