Purification of an L-amino acid oxidase from Bungarus caeruleus (Indian krait) venom

• Main Authors: SS More, KM Kiran, SM Veena, JR Gadag
• Format: Article in Journal/Newspaper
• Language: English
• Published: SciELO 2010
• Subjects: L-amino acid oxidase; Bungarus caeruleus; platelet aggregation; Arctic medicine. Tropical medicine; RC955-962; Toxicology. Poisons; RA1190-1270; Zoology; QL1-991; Arctic; Indian
• Online Access: https://doaj.org/article/c0ce68dd7ccc40928f100a19dfc48a6d

Snake venoms are rich in enzymes such as phospholipase A2, proteolytic enzymes, hyaluronidases and phosphodiesterases, which are well characterized. However, L-amino acid oxidase (LAO EC.1.4.3.2) from snake venoms has not been extensively studied. A novel L-amino acid oxidase from Bungarus caeruleus venom was purified to homogeneity using a combination of ion-exchange by DEAE-cellulose chromatography and gel filtration on Sephadex® G-100 column. The purified monomer of LAO showed a molecular mass of 55 ±1 kDa estimated by SDS-PAGE. The specific activity of purified LAO was 6,230 ± 178 U/min/mg, versus 230 ± 3.0 U/min/mg for the whole desiccated venom, suggesting a 27-fold purification with a 25% yield. Optimal pH and temperature for maximum purified enzyme activity were 6.5 and 37ºC, respectively. Platelet aggregation studies show that purified LAO inhibited ADP-induced platelet aggregation dose-dependently at 0.01 to 0.1 µM with 50% inhibitory concentration (IC50) of 0.04 µM, whereas at a 0.08 µM concentration it did not induce appreciable aggregation on normal platelet-rich plasma (PRP). The purified protein catalyzed oxidative deamination of L-amino acids while the most specific substrate was L-leucine. The purified LAO oxidizes only L-forms, but not D-forms of amino acids, to produce H2O2. The enzyme is important for the purification and determination of certain amino acids and for the preparation of α-keto acids.