Identification of the enantiomeric nature of 2-keto-3-deoxy-galactonate in the catabolic pathway of 3,6-anhydro-l-galactose

A novel metabolic pathway of 3,6-anhydro- l -galactose ( l -AHG), the main sugar component in red macroalgae, was first discovered in the marine bacterium Vibrio sp. EJY3. l -AHG is converted to 2-keto-3-deoxy-galactonate (KDGal) in two metabolic steps. Here, we identified the enantiomeric nature of KDGal in the l -AHG catabolic pathway via stereospecific enzymatic reactions accompanying the biosynthesis of enantiopure l -KDGal and d -KDGal. Enantiopure l -KDGal and d -KDGal were synthesized by enzymatic reactions derived from the fungal galacturonate and bacterial oxidative galactose pathways, respectively. KDGal, which is involved in the l -AHG pathway, was also prepared. The results obtained from the reactions with an l -KDGal aldolase, specifically acting on l -KDGal, showed that KDGal in the l -AHG pathway exists in an l -enantiomeric form. Notably, we demonstrated the utilization of l -KDGal by Escherichia coli for the first time. E. coli cannot utilize l -KDGal as the sole carbon source. However, when a mixture of l -KDGal and d -galacturonate was used, E. coli utilized both. Our study suggests a stereoselective method to determine the absolute configuration of a compound. In addition, our results can be used to explore the novel l -KDGal catabolic pathway in E. coli and to construct an engineered microbial platform that assimilates l -AHG or l -KDGal as substrates. Key points • Stereospecific enzyme reactions were used to identify enantiomeric nature of KDGal • KDGal in the l -AHG catabolic pathway exists in an l -enantiomeric form • E. coli can utilize l -KDGal as a carbon source when supplied with d -galacturonate