Biochemical Characteristics and Substrate Degradation Pattern of a Novel Exo-Type β-Agarase from the Polysaccharide-Degrading Marine Bacterium Flammeovirga sp. Strain MY04

Abstract

Exo-type agarases can degrade agarose to yield disaccharides almost exclusively, and therefore, they are important tools for disaccharide preparation. However, their enzymatic mechanisms and agarose degradation patterns are still unclear due to the lack of proper technologies for sugar chain analysis. In this study, AgaO was identified as an exo-type agarase of agarose-degrading Flammeovirga bacteria, representing a novel branch of glycoside hydrolase family 50. Using fluorescence labeling, high-performance liquid chromatography, and mass spectrum analysis technologies, we provide a useful oligosaccharide sequencing method to determine the patterns of enzyme cleavage of glycosidic bonds. We also demonstrate that AgaO produces neoagarobiose by gradually cleaving disaccharides from the nonreducing end of fluorescently labeled sugars. This study will benefit future enzyme applications and oligosaccharide studies.