Engineering Embden-Meyerhof-Parnas Glycolysis to Generate Noncanonical Reducing Power.

ABSTRACT: Noncanonical cofactors such as nicotinamide mononucleotide (NMN⁺) supplant the electron-transfer functionality of the natural cofactors, NAD(P)⁺, at a lower cost in cell-free biomanufacturing and enable orthogonal electron delivery in whole-cell metabolic engineering. Here, we redesign the high-flux Embden-Meyerhof-Parnas (EMP) glycolytic pathway to generate NMN⁺-based reducing power, by engineering Streptococcus mutans glyceraldehyde-3-phosphate dehydrogenase (Sm GapN) to utilize NMN⁺. Through iterative rounds of rational design, we discover the variant GapN Penta (P179K-F153S-S330R-I234E-G210Q) with high NMN⁺-dependent activity and GapN Ortho (P179K-F153S-S330R-I234E-G214E) with ~3.4 × 10⁶-fold switch in cofa

SUBMITTER: King E 

PROVIDER: S-EPMC10449333 | biostudies-literature | 2022 Jul

REPOSITORIES: biostudies-literature