Exploring the substrate scope of glycerol dehydrogenase GldA from E. coli BW25113 towards cis-dihydrocatechol derivatives

Glycerol dehydrogenase (GldA) from Escherichia coli BW25113, naturally catalyzes the oxidation of glycerol to dihydroxyacetone. It is known that GldA exhibits promiscuity towards short-chain C2-C4 alcohols. However, there are no reports regarding the substrate scope of GldA towards larger substrates. Herein we demonstrate that GldA can accept bulkier C6-C8 alcohols than previously anticipated. Overexpression of the gldA gene in the knockout background, E. coli BW25113 Delta gldA, was strikingly effective converting 2 mM of the compounds: cis- dihydrocatetechol, cis-(1 S,2 R)-3-methylcyclohexa-3,5-diene-1,2-diol and cis-(1 S,2 R)- 3-ethylcyclohexa-3,5-diene-1,2-diol, into 2.04 +/- 0.21 mM of catechol, 0.62 +/- 0.11 mM 3-methylcatechol, and 0.16 +/- 0.02 mM 3-ethyl-catechol, respectively. In-silico studies on the active site of GldA enlightened the decrease in product formation as the steric substrate demand increased. These results are of high interests for E. coli-based cell factories expressing Rieske non-heme iron dioxygenases, producing cis-dihydrocatechols, since such sough-after valuable products can be immediately degraded by GldA, substantially hampering the expected performance of the recombinant platform.

Automated summary

Glycerol dehydrogenase (GldA) from Escherichia coli can oxidize glycerol to dihydroxyacetone and accept larger C6-C8 alcohols as substrates. This is important for E. coli-based cell factories expressing Rieske non-heme iron dioxygenases, as GldA can degrade valuable products, hampering the expected performance of the recombinant platform.