Chemical modification of Lys-6 in Taiwan cobra phospholipase A2 with 4-chloro-3,5-dinitrobenzoate.
Phospholipase A2 (PLA2) from Naja naja atra snake venom was modified with 4-chloro-3,5-dinitrobenzoate, and one major carboxydinitrophenylated (CDNP) PLA2 was separated by high performance liquid chromato-graphy. CDNP-PLA2 contained only one CDNP group on Lys-6 and showed a 93% drop in enzymatic activity. However, carboxydinitrophenylation did not significantly affect the secondary structure of the enzyme molecule as revealed by the CD spectra, and Ca2+ binding and antigenicity of CDNP-PLA2 were unaffected. Conversion of nitro groups to amino groups resulted in a partial restoration of enzymatic activity of CDNP-PLA2 to 35% of that of native enzyme. These results suggested that the positively charged side chain of Lys-6 played a role in the enzymatic mechanism of PLA2. However, the partial restoration in PLA2 activity reflects that a distortion of the active conformation arising from incorporation of a bulky CDNP group should occur.
Chang
L S
LS
Department of Biochemistry, Kaohsiung Medical College, Taiwan, ROC.
Hung
J J
JJ
Lin
S
S
Chang
C C
CC
D000595:Amino Acid Sequence; D000818:Animals; D000941:Antigens; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism; D002723:Chlorobenzoates / Q000494:pharmacology*; D002942:Circular Dichroism; D004546:Elapid Venoms; D017815:Elapidae; D007202:Indicators and Reagents; D007700:Kinetics; D008239:Lysine; D010741:Phospholipases A / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism*; D054467:Phospholipases A2; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D013624:Taiwan
D000941:Antigens; D002723:Chlorobenzoates; D004546:Elapid Venoms; D007202:Indicators and Reagents; C015746:4-chloro-3,5-dinitrobenzoate; D010741:Phospholipases A; D054467:Phospholipases A2; D008239:Lysine; D002118:Calcium