ArticleMicrobial biotechnology2025
Reconstitution of Methionine Cycle With ATP Regeneration for Whole-Cell Catalysis of Creatine Production in Engineered Escherichia coli.
Article in Microbial biotechnology, 2025. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 3 papers.
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3 citing papers in PubMed.
- Toward engineered microbial chassis for growth-production balancing.Synthetic and systems biotechnology · 2027Review
- Synthesis and Biological Activity Characterization of Vascular Endothelial Growth Factor Using an Optimized Wheat Germ Cell-Free System.Current issues in molecular biology · 2026Article
- Dual-Enzyme Co-Catalysis Strategy for Fructooligosaccharides (FOS) Biocatalytic Synthesis for Valorization of Low-Cost Byproduct Sugarcane Molasses.Foods (Basel, Switzerland) · 2026Article
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9 authors.
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Abstract
Creatine (CR) is a naturally occurring amino acid derivative that plays a key role in cellular energy homeostasis, which has wide-ranging applications in food and medicine. Currently, the lack of green and sustainable CR biomanufacturing methods has led to reliance on chemical methods for industrial CR synthesis. This study presents a biological approach to synthesising CR using whole-cell catalysis by engineered Escherichia coli. First, through screening of critical enzymes from different sources and dual-enzyme co-expression strategies, arginine: glycine amidinotransferase (AGAT) from Amycolatopsis kentuckyensis and guanidinoacetate N-methyltransferase (GAMT) from Mus caroli were introduced to construct the CR biosynthesis pathway, yielding 0.83 g/L CR production. Then, the expression level of GAMT, the critical rate-limiting enzyme, was optimised by screening the ribosome binding site and N-terminal coding sequences, resulting in a 92% enhancement of CR production, reaching 1.59 g/L. Next, the endogenous ornithine and methionine cycles were further engineered to boost the synthesis of the precursor guanidinoacetate (GAA) and methyl donor S-adenosylmethionine (SAM), leading to a 68% increase in CR production, reaching 2.67 g/L. Finally, considering adenosine triphosphate (ATP) is required as a cofactor for SAM biosynthesis, we integrated the reconstitution methionine cycle with a polyphosphate kinase-based ATP regeneration system, achieving a CR titre of 5.27 g/L with a productivity of 0.22 g/L/h, and the molar conversion of substrate arginine was 71 mol% over 24 h following the engineering process. This study is the first report achieving whole-cell catalysis of CR production in engineered E. coli with a dual enzyme cascade using arginine as substrate, providing a new platform for CR production and insights into the biosynthesis of high-value metabolites that rely on ATP consumption.
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