NADP+-dependent dehydrogenase SCO3486 and cycloisomerase SCO3480 are key enzymes for 3,6-anhydro-L-galactose catabolism in Streptomyces coelicolor A3(2).

Agarose is a linear polysaccharide composed of d-galactose and 3,6-anhydro-l-galactose. It is a major component of the red algal cell wall, and is gaining attention as an abundant marine biomass. However, the inability to ferment 3,6-anhydro-l-galactose is considered an obstacle in the large-scale use of agarose, which could be addressed by understanding 3,6-anhydro-l-galactose catabolism in agarolytic microorganisms. Since 3,6-anhydro-l-galactose catabolism was uniquely confirmed in Vibrio sp. EJY3, a gram-negative marine bacterial species, we investigated AHG metabolism in Streptomyces coelicolor A3 (2), an agarolytic gram-positive soil bacterium. Based on genomic data, the SCO3486 protein (492 amino acids) and the SCO3480 protein (361 amino acids) of S. coelicolor A3 (2) showed identity with H2IFE7.1 (40% identity) encoding 3,6-anhydro-l-galactose dehydrogenase and H2IFX0.1 (42% identity) encoding 3,6-anhydro-l-galactonate cycloisomerase, respectively, which are involved in the initial catabolism of 3,6-anhydro-l-galactose in Vibrio sp. EJY3. Thin layer chromatography and mass spectrometry of the bioconversion products catalyzed by recombinant SCO3486 and SCO3480 proteins, revealed that SCO3486 is an 3,6-anhydro-l-galactose dehydrogenase that oxidizes 3,6-anhydro-l-galactose to 3,6-anhydro-l-galactonate, and SCO3480 is a 3,6-anhydro-l-galactonate cycloisomerase that converts 3,6-anhydro-l-galactonate to 2-keto-3-deoxygalactonate. SCO3486 showed maximum activity at pH 6.0 at 50°C, increased activity in presence of iron ions, and activity against various aldehyde substrates, which is quite distinct from 3,6-anhydro-l-galactose-specific H2IFE7.1 in Vibrio sp. EJY3. Therefore, the catabolic pathway of 3,6-anhydro-L-galactose seems to be similar in most agar-degrading microorganisms, but the enzymes involved appear to be very diverse.