Antioxidant and antidiabetic activities of vanadium binding proteins purified from the sea squirt Halocynthia roretzi

Sea squirts accumulate vanadium compounds with potent antidiabetic activity, which are involved in immune defense. In this study, vanadium concentrations of fresh blood plasma, intestine, and muscle of the sea squirt Halocynthia roretzi were 6.3, 3.7 and 2.1 mg/kg respectively. Two vanadium binding proteins (VBPs) from blood plasma and intestine were purified through (NH₄)₂SO₄ precipitation, and DEAE-Sepharose ion exchange and Sephacryl S-200 HR gel filtration chromatography, in that order. The purity and yield of the intestine and blood plasma vanadium binding proteins, VBP_intestine and VBP_{blood plasma,} were 13.4 folds and 7.1%, and 20.9 folds and 6.8%, respectively. There were two protein bands on the sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) with molecular weights of 24.3 and 68.8 kDa and one with 96.7 kDa on the native-PAGE of VBP_{blood plasma}, whereas only one protein band of VBP_intestine on the SDS-PAGE with 26.5 kDa. Antioxidant activities of VBPs were lower than that of ascorbic acid. Both VBPs exerted strong inhibitory activity against Saccharomyces cerevisiae and mild against Bacillus stearothermophilus and rat intestinal α-glucosidase. IC₅₀ values of VBP_intestine and VBP_{blood plasma} against S. cerevisiae α‐glucosidase were 28.34 and 12.60 μg/ml, respectively. The K_m, V_max, k_cat, and k_cat/K_m values of VBP_intestine and VBP_{blood plasma} were 4.29, 0.036, 6.58 and 1.53 × 10³, and 7.63 mM, 0.057 mM/min, 10.41 s⁻¹ and 1.36 × 10³ (M sec)⁻¹, respectively. There was a synergistic interaction between VBP_{blood plasma} and VBP_intestine on rat intestinal α-glucosidase inhibitory activity.