A Novel Agarase, Gaa16B, Isolated from the Marine Bacterium <i>Gilvimarinus agarilyticus</i> JEA5, and the Moisturizing Effect of Its Partial Hydrolysis Products

We recently identified a <inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><mi mathvariant="sans-serif">β</mi></semantics></math></inline-formula>-agarase, Gaa16B, in the marine bacterium <i>Gilvimarinus agarilyticus</i> JEA5. Gaa16B, belonging to the glycoside hydrolase 16 family of <inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><mi mathvariant="sans-serif">β</mi></semantics></math></inline-formula>-agarases, shows less than 70.9% amino acid similarity with previously characterized agarases. Recombinant Gaa16B lacking the carbohydrate-binding region (rGaa16Bc) was overexpressed in <i>Escherichia coli</i> and purified. Activity assays revealed the optimal temperature and pH of rGaa16Bc to be 55 <inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><msup><mrow></mrow><mo>∘</mo></msup></semantics></math></inline-formula>C and pH 6–7, respectively, and the protein was highly stable at 55 <inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><msup><mrow></mrow><mo>∘</mo></msup></semantics></math></inline-formula>C for 90 min. Additionally, rGaa16Bc activity was strongly enhanced (2.3-fold) in the presence of 2.5 mM MnCl₂. The <i>K_m</i> and <i>V_max</i> of rGaa16Bc for agarose were 6.4 mg/mL and 953 U/mg, respectively. Thin-layer chromatography analysis revealed that rGaa16Bc can hydrolyze agarose into neoagarotetraose and neoagarobiose. Partial hydrolysis products (PHPs) of rGaa16Bc had an average molecular weight of 88–102 kDa and exhibited > 60% hyaluronidase inhibition activity at a concentration of 1 mg/mL, whereas the completely hydrolyzed product (CHP) showed no hyaluronidase at the same concentration. The biochemical properties of Gaa16B suggest that it could be useful for producing functional neoagaro-oligosaccharides. Additionally, the PHP of rGaa16Bc may be useful in promoting its utilization, which is limited due to the gel strength of agar.