Purification and characterization of phospholipase A2 from Trimeresurus gramineus venom.
Phospholipase A2 was purified to homogeneity from the venom of Trimeresurus gramineus (the Green Habu snake) via three steps consisting of Sephadex G-75, DEAE-cellulose, and DEAE-Toyopearl 650M column chromatographies. Molecular weight determinations showed that the enzyme consists of a single polypeptide chain with a molecular weight of approximately 14,000. The isoelectric point was 4.5. The enzyme is characterized by high contents of acidic amino acids, glycine, and half-cystine. Calcium ion was essential for eliciting activity. The enzyme was inactivated by alkylation of a single histidine residue with p-bromophenacyl bromide following pseudo first-order kinetics, and the rate of the inactivation was depressed in the presence of Ca2+. The N-terminal sequence of this enzyme determined to the 40th residue was found to be highly homologous to that of Trimeresurus okinavensis phospholipase A2 but not to that of Trimeresurus flavoviridis phospholipase A2. The phenylalanine residue at the 27th position of T. gramineus phospholipase A2 is noteworthy because all other phospholipases A2, with only two exceptions, contain a tyrosine residue at this position.
Oda
N
N
Department of Chemistry, Faculty of Science, Kyushu University, Fukuoka.
Sakai
H
H
Shieh
T C
TC
Nakamura
H
H
Sakamoto
S
S
Kihara
H
H
Chang
C C
CC
Ohno
M
M
D000098:Acetophenones / Q000494:pharmacology; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D002118:Calcium / Q000494:pharmacology; D002413:Cations, Divalent; D002845:Chromatography; D003435:Crotalid Venoms / Q000032:analysis*; D004789:Enzyme Activation / Q000187:drug effects; D007526:Isoelectric Point; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments; D010713:Phosphatidylcholines / Q000378:metabolism; D010740:Phospholipases / Q000302:isolation & purification*; D010741:Phospholipases A / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D054467:Phospholipases A2
D000098:Acetophenones; D000596:Amino Acids; D002413:Cations, Divalent; D003435:Crotalid Venoms; D010446:Peptide Fragments; D010713:Phosphatidylcholines; C029748:Trimeresurus venoms; D010740:Phospholipases; D010741:Phospholipases A; D054467:Phospholipases A2; C006821:4-bromophenacyl bromide; D002118:Calcium; C028694:1-palmitoyl-2-oleoylphosphatidylcholine