title: Amino acid sequence of a basic Agkistrodon halys blomhoffii phospholipase A2. Possible role of NH2-terminal lysines in action on phospholipids of Escherichia coli.
abstract: A basic (pI = 10.2) phospholipase A2 of the venom of the snake Agkistrodon halys blomhoffii is one of a few phospholipases A2 capable of hydrolyzing the phospholipids of Escherichia coli killed by a bactericidal protein purified from human or rabbit neutrophil granules. We have shown that modification of as many as 4 mol of lysine per mole of the phospholipase A2, either by carbamylation or by reductive methylation [Forst, S., Weiss, J., & Elsbach, P. (1982) J. Biol. Chem. 257, 14055-14057], had no effect on catalytic activity toward extracted E. coli phospholipids or the phospholipids of autoclaved E. coli. In contrast, modification of 1 mol of lysine per mole of enzyme substantially reduced activity toward the phospholipids of E. coli killed by the neutrophil protein. To explore further the role of lysines in the function of this phospholipase A2, we determined the amino acid sequence of the enzyme and the incorporation of [14C]cyanate into individual lysines when, on average, 1 lysine per molecule of enzyme had been carbamylated. After incorporation of approximately 1 mol of [14C]cyanate per mole of protein, the phospholipase A2 was reduced, alkylated, and exhaustively carbamylated with unlabeled cyanate. The amino acid sequence was determined of the NH2-terminal 33 amino acids of the holoprotein and of peptides isolated after digestion with trypsin and Staphylococcus aureus V-8 protease. The protein contains 122 amino acid residues, 17 of which are lysines. The NH2-terminal region is unique among more than 30 phospholipases A2 previously sequenced because of its high content of basic residues (His-1, Arg-6, and Lys-7, -10, -11, and -15).(ABSTRACT TRUNCATED AT 250 WORDS)
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D003435:Crotalid Venoms / Q000378:metabolism*; D010450:Endopeptidases; D004926:Escherichia coli; D007700:Kinetics; D010446:Peptide Fragments / Q000032:analysis; D010740:Phospholipases / Q000378:metabolism*; D010741:Phospholipases A / Q000302:isolation & purification / Q000378:metabolism*; D054467:Phospholipases A2; D010743:Phospholipids / Q000378:metabolism; D011487:Protein Conformation; D012697:Serine Endopeptidases; D013379:Substrate Specificity; D014357:Trypsin
chemical_list: C031442:Agkistrodon venoms; D003435:Crotalid Venoms; D010446:Peptide Fragments; D010743:Phospholipids; D010740:Phospholipases; D010741:Phospholipases A; D054467:Phospholipases A2; D010450:Endopeptidases; D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase; D014357:Trypsin