Overexpression of pyrroloquinoline quinone biosynthetic genes affects l-sorbose production in Gluconobacter oxydans WSH-003

• Overexpression of each single gene enhanced PQQ biosynthesis except the tldD.
• The tufB promoter is beneficial for overexpressing the PQQ biosynthetic genes.
• PqqB protein is the key enzyme that transports PQQ across the membrane.
• Higher PQQ level facilitated the conversion of d-sorbitol to l-sorbose.

Pyrroloquinoline quinone (PQQ) is a cofactor of various membrane-bound dehydrogenases. The amount of endogenous PQQ is generally regarded as a bottleneck to achieving higher catalytic efficiency of PQQ-dependent dehydrogenases. Proteins that biosynthesize PQQ in Gluconobacter oxydans WSH-003 are encoded by the pqqABCDE gene cluster and the tldD gene. In this study, PQQ overproduction in G. oxydans was attempted by overexpressing PQQ biosynthetic genes using the promoter of pqqA and the elongation factor TU (tufB). Overexpression of each single gene could enhance PQQ biosynthesis except for the tldD gene. Overexpression of pqqA, pqqB, pqqC, pqqD and pqqE with the pqqA promoter enhanced the extracellular PQQ concentration by 38.5%, 68.4%, 19.9%, 30.3% and 8.2%, respectively, whereas production was enhanced by 59.4%, 85.7%, 30.9%, 42.2% and 19.3%, respectively, using the tufB promoter. In addition, the results show that PQQ biosynthesis could be enhanced by overexpressing some of the individual genes in the gene cluster in G. oxydans and the PQQ levels were positively correlated with the efficiency of conversion of d-sorbitol to l-sorbose. The results demonstrated that cofactor engineering of PQQ in G. oxydans is beneficial for enhancing the production of quinoprotein-related products.

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