Met-8 of the beta 1-bungarotoxin phospholipase A2 subunit is essential for the phospholipase A2-independent neurotoxic effect.
beta 1-Bungarotoxin consists of a phospholipase A2 subunit and a non-phospholipase A2 subunit. The toxin was oxidized with a 100-fold molar excess of chloramine T with respect to the methionine content of the protein in 0.1 M Tris/HCl at pH 8.5 and at room temperature. Reactivities of the two methionine (Met-6 and Met-8 of the phospholipase A2 subunit), five histidine, 14 tyrosine and one tryptophan residues of one toxin molecule with chloramine T were assessed from the change in intrinsic fluorescence and amino acid composition of the protein. Met-8 and one tyrosine on the phospholipase A2 subunit and less than one histidine were oxidized, while Met-6 remained intact after 30 min of reaction. One histidine and approx. two tyrosine residues were oxidized when both methionine residues were oxidized after 90 min of reaction. The sole tryptophan was oxidized slightly throughout the reaction. The chloramine T oxidation did not destroy the two Ca(2+)-binding domains, though it modified the toxin to become less effective at binding Ca2+. The modified toxin obtained after 30 or 90 min reaction time retained 65% or 40% of the phospholipase A2 activity of the parent toxin, but both were not lethal to mice and showed a very weak ability to induce the indirectly evoked contraction of chick biventer cervicis muscle. It is suggested that Met-8 may play an important role in the phospholipase A2-independent interaction with the nerve terminal membrane during the neurotoxic effect of beta 1-bungarotoxin.
Chu
S T
ST
Institute of Biological Chemistry, Academia Sinica, Taipei, Taiwan, Republic of China.
Chu
C C
CC
Tseng
C C
CC
Chen
Y H
YH
D000818:Animals; D001665:Binding Sites; D002038:Bungarotoxins / Q000737:chemistry / Q000378:metabolism / Q000633:toxicity*; D002118:Calcium / Q000378:metabolism; D002645:Chickens; D002700:Chloramines / Q000378:metabolism; D006639:Histidine / Q000378:metabolism; D008715:Methionine / Q000378:metabolism; D009119:Muscle Contraction; D009132:Muscles / Q000294:innervation / Q000502:physiology; D010084:Oxidation-Reduction; D010741:Phospholipases A / Q000378:metabolism*; D054467:Phospholipases A2; D013050:Spectrometry, Fluorescence; D013329:Structure-Activity Relationship; D014105:Tosyl Compounds / Q000378:metabolism; D014364:Tryptophan / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism
D002038:Bungarotoxins; D002700:Chloramines; D014105:Tosyl Compounds; C016300:chloramine-T; D014443:Tyrosine; D006639:Histidine; D014364:Tryptophan; D008715:Methionine; D010741:Phospholipases A; D054467:Phospholipases A2; D002118:Calcium