Chemical modification of Lys-6 in phospholipase A2 from Naja melanoleuca snake venom.
Phospholipase A2 isolated from Naja melanoleuca snake venom was modified with 4-chloro-3,5-dinitrobenzoate. The modification reaction was accelerated by the cofactor Ca2+. Micellar concentrations of the substrate analog n-tetradecylphosphocholine retarded the inactivation of the enzyme. Modification resulted in the incorporation of one mole dinitrobenzoate/mole of protein. The modified residue could be identified as Lys-6 in the amino acid sequence of the enzyme. Although the modified enzyme still retains a high affinity for micellar substrate analogs and the cofactor Ca2+, it had lost nearly all its activity towards micellar substrates. Upon reduction of the nitro groups in the modified protein the enzymatic activity could be restored by more than 50%. Therefore Lys-6 cannot be directly involved in enzymatic catalysis. It is concluded that introduction of the negatively charged dinitrobenzoate group on Lys-6 gives rise to a distortion of the active site of the enzyme.
D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism; D002384:Catalysis; D055598:Chemical Phenomena; D002621:Chemistry; D004546:Elapid Venoms; D008239:Lysine; D010740:Phospholipases / Q000378:metabolism; D010741:Phospholipases A / Q000378:metabolism; D054467:Phospholipases A2; D011485:Protein Binding; D013329:Structure-Activity Relationship
D004546:Elapid Venoms; D010740:Phospholipases; D010741:Phospholipases A; D054467:Phospholipases A2; D008239:Lysine; D002118:Calcium