Correlation between the phospholipids domains of the target cell membrane and the extent of Naja kaouthia PLA(2)-induced membrane damage: evidence of distinct catalytic and cytotoxic sites in PLA(2) molecules.
Two phospholipase A(2) (PLA(2)) enzymes (NK-PLA(2)-A and NK-PLA(2)-B) were purified from the venom of the monocled cobra Naja kaouthia. The molecular weights of NK-PLA(2)-A and NK-PLA(2)-B, as estimated by mass spectrometry, were 13,619 and 13,303 Da respectively. Both phospholipases were highly thermostable, had maximum catalytic activity at basic pH, and showed preferential hydrolysis of phosphatidylcholine. Intravenous injection of either PLA(2) up to a dose of 10 mg/kg body weight was non-toxic to mice and did not show neurotoxic symptoms. The N. kaouthia PLA(2)s displayed anticoagulant and cytotoxic activity, but poor hemolytic activity. Both the PLA(2)s were more toxic to Sf9 and Tn cells compared to VERO cells. NK-PLA(2) exhibited selective lysis of wild-type baculovirus-infected Sf9 cells compared to normal cells. Amino acid modification studies and heating experiments suggest that separate sites in the NK-PLA(2) molecules are responsible for their catalytic, anticoagulant and cytotoxic activities.
Mukherjee
Ashis K
AK
Department of Molecular Biology and Biotechnology, Tezpur University, Tezpur-784 028, Assam, India. akm@tezu.ernet.in
D000328:Adult; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D000925:Anticoagulants / Q000494:pharmacology; D001665:Binding Sites; D020134:Catalytic Domain; D002460:Cell Line; D002522:Chlorocebus aethiops; D017815:Elapidae; D004926:Escherichia coli / Q000187:drug effects; D006023:Glycoproteins / Q000633:toxicity*; D006461:Hemolysis; D006801:Humans; D007700:Kinetics; D008563:Membrane Lipids / Q000378:metabolism*; D008970:Molecular Weight; D010741:Phospholipases A / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D010743:Phospholipids / Q000378:metabolism*; D017433:Protein Structure, Secondary; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D018411:Spodoptera; D014162:Transfection; D014709:Vero Cells
D000900:Anti-Bacterial Agents; D000925:Anticoagulants; D006023:Glycoproteins; D008563:Membrane Lipids; D010743:Phospholipids; C090541:phospholipase A2 inhibitory protein, Naja naja kaouthia; D010741:Phospholipases A