Discovery of the Novel Metabolic Pathway of 3,6-Anhydro-L-galactose, the Main Sugar of Red Macroalgae by the Integrative Analyses of Transcriptome and Metabolome

Recently, red macroalgae are being actively investigated as the carbohydrate resource for producing bio-based products. The main carbohydrate composed of red macroalgae is agarose, which is a heteropolysaccharide composed of D-galactose and 3,6-anhydro-L-galactose (AHG). Although AHG is the major sugar in red macroalgae, the metabolic fate of AHG is completely unknown in any organisms. AHG is not catabolized by ordinary fermentative microorganisms, and it hampers the utilization of red macroalgae as renewable biomass for biofuel and chemical production. In this study, we have discovered the novel metabolic pathway of AHG in a marine microorganism by integrative analyses of metabolome and transcriptome. Using the Vibrio sp. strain EJY3 as a model organism, we performed metabolite and transcriptomic analyses to identify key metabolic intermediates and the corresponding enzymes that are possibly involved in AHG pathway. The analyses of metabolome and transcriptome of EJY3 revealed two key metabolic intermediates of AHG, 3,6-anhydrogalactonate (AHGA) and 2-keto-3-deoxy-galactonate. The oxidation by an NA(D)P⁺‒dependent AHG dehydrogenase and isomerization by an AHGA cycloisomerase are the key metabolic steps for metabolizing AHG. This newly discovered metabolic route was also verified in vivo by demonstrating the growth of E. coli harboring the genes of these two enzymes on AHG as a sole carbon source. This is the first report on the discovery of the novel metabolic pathway of AHG in a living organism by systematic integrative analyses of transcriptome and metabolome.