Rare sugar production by coupling of NADH oxidase and l-arabinitol dehydrogenase

Abstract

An efficient biocatalytic cell-free system containing L-arabinitol dehydrogenase (LAD) for L-arabinitol oxidation and NADH oxidase (Nox) for cofactor regeneration was successfully constructed and used for L-rare sugar production. The recombinant LAD (HjLAD) from Hypocrea jecorina suffered from NADH inhibition. Applying Nox to the cell-free HjLAD system drove the thermodynamically unfavorable equilibrium from L-arabinitol to L-xylulose, along with the regeneration of the oxidized cofactor NAD + . Response surface methodology was used to optimize the conditions for L-xylulose production. The optimal conditions for biocatalytic production of L-xylulose were found to be 4.9 U/mL HjLAD, 8.2 mM NAD + at pH 8.0, and 30.9 o C; a high conversion rate of 78.4% was achieved under these conditions. We demonstrate a cellfree enzyme coupling system enabling efficient cofactor recycling and providing high yields of L-xylulose. The results indicate that this coupling system provides a new biocatalytic method for L-rare sugar production.

Protein–inorganic hybrid system for efficient his-tagged enzymes immobilization and its application in l-xylulose production

“…Thus, L-xylulose production was evaluated at pH 8.0. 29 The free enzyme combination of HjLAD and SpNox resulted in the maximum conversion of L-arabinitol to Lxylulose with a yield of 31.2% aer incubation for 6 h under optimum conditions. In contrast, immobilized HjLAD and SpNox enzymes nanoowers showed 2.9-fold higher conversion with a maximum yield of 90.1%.…”

“…Rare sugars are unique monosaccharides that play a signicant role as recognition elements in bio-active molecules with a broad range of applications. [29][30][31] L-Xylulose is a rare sugar used primarily as a potential inhibitor of a-glucosidases and indicator of liver cirrhosis disease. 32 The conversion of carbohydrates by metal catalysts are known due their high selectivity and stability.…”

Overcoming NADPH product inhibition improves D-sorbitol conversion to L-sorbose

“…Cofactor concentration is another important parameter in the catalytic process and requires careful monitoring because high cofactor concentrations may lead to enzyme inhibition and high production costs 18,19,28 . The effect of cofactor concentration on l -sorbose production was investigated (Fig.…”

Engineering Spatially Organized Multienzyme Assemblies for Complex Chemical Transformation

“…When cofactor-dependent enzymes cannot be avoided, cofactors must be regenerated in situ for the process to be economically feasible. Several regeneration strategies have been attempted including electrochemical and chemical regeneration systems, cofactor immobilization, and supplementing the reaction with additional enzymes for enzymatic cofactor regeneration. − Of all these strategies, enzymatic cofactor regeneration schemes have been the most-studied, and several types of enzymes have been used to regenerate nicotinamide cofactors. For example, NAD­(P)H can be regenerated from NAD­(P) by adding alcohol dehydrogenase, formate dehydrogenase, or glucose dehydrogenase along with its respective sacrificial substrate to a reaction in which NAD­(P)­H is oxidized.…”