One-Pot Bioconversion of l-Arabinose to l-Ribulose in an Enzymatic Cascade

Abstract

This work reports the one-pot enzymatic cascade that completely converts l-arabinose to l-ribulose using four reactions catalyzed by pyranose 2-oxidase (P2O), xylose reductase, formate dehydrogenase, and catalase. As wild-type P2O is specific for the oxidation of six-carbon sugars, a pool of P2O variants was generated based on rational design to change the specificity of the enzyme towards the oxidation of l-arabinose at the C2-position. The variant T169G was identified as the best candidate, and this had an approximately 40-fold higher rate constant for the flavin reduction (sugar oxidation) step, as compared to the wild-type enzyme. Computational calculations using quantum mechanics/molecular mechanics (QM/MM) molecular dynamics (MD) showed that this improvement is due to a decrease in the steric effects at the axial C4-OH of l-arabinose, which allows a reduction in the distance between the C2-H and flavin N5, facilitating hydride transfer and enabling flavin reduction.

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Recommended citation: Chuaboon L., Wongnate T., Punthong P., Kiattisewee C., Lawan N., Hsu C.Y., Lin C.H., Bornscheuer U.T., Chaiyen P.^† (2019). "One-Pot Bioconversion of l-Arabinose to l-Ribulose in an Enzymatic Cascade." Angewandte Chemie International Edition. 58(8):2428-2432. PMID: 30605256.