Histidine and lysine residues and the activity of phospholipase A2 from the venom of Bitis gabonica.
Chemical modification of phospholipase A2 (phosphatide 2-acyl-hydrolase, EC 3.1.1.4) from the venom of gaboon adder (Bitis gabonica) showed that histidine and lysine residues are essential for enzyme activity. Treatment with p-bromophenacyl bromide or pyridoxal 5'-phosphate resulted in the specific covalent modification of one histidine or a total of one lysine residue per molecule of enzyme, respectively, with a concomitant loss of enzyme activity. Competitive protection against modification and inactivation was afforded by the presence of Ca2+ and/or micellar concentrations of substrate analogue, lysophosphatidylcholine. Neither modification caused any significant conformational change, as judged from circular dichroic properties. Amino acid analyses and the alignment of peptides from cyanogen bromide and proteolytic cleavage of modified enzyme preparations delineated His-45 as the only residue modified by p-bromophenacyl bromide. However, pyridoxal 5'-phosphate was shown to have reacted not with a single lysine but with four different ones (residues 11, 33, 58 and 111) in such a manner that an overall stoichiometry of one modified lysine residue/molecule enzyme resulted. Apparently, the essential function of lysine could be fulfilled by any one out of these four residues.
D000098:Acetophenones / Q000494:pharmacology; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D002118:Calcium / Q000494:pharmacology; D002942:Circular Dichroism; D006639:Histidine; D006863:Hydrogen-Ion Concentration; D008239:Lysine; D008244:Lysophosphatidylcholines / Q000494:pharmacology; D010446:Peptide Fragments / Q000032:analysis; D010740:Phospholipases / Q000037:antagonists & inhibitors*; D011732:Pyridoxal Phosphate / Q000494:pharmacology; D012910:Snake Venoms; D013329:Structure-Activity Relationship
D000098:Acetophenones; D000596:Amino Acids; D008244:Lysophosphatidylcholines; D010446:Peptide Fragments; D012910:Snake Venoms; D006639:Histidine; D011732:Pyridoxal Phosphate; D010740:Phospholipases; D008239:Lysine; C006821:4-bromophenacyl bromide; D002118:Calcium