Purification and characterization of a novel glutathione S-transferase from Atactodea striata.
A novel GST isoenzyme was purified from hepatopancreas cytosol of Atactodea striata with a combination of affinity chromatography and reverse-phase HPLC. The molecular weight of the enzyme was determined to be 24 kDa by SDS-PAGE electrophoresis and 48 kDa by gel chromatography, in combination with GST information from literature revealed that the native enzyme was homodimeric with a subunit of M(r) 24 kDa. The purified enzyme, exhibited high activity towards 1-chloro-2,4-dinitrobenzene (CDNB) and 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl). Kinetic analysis with respect to CDNB as substrate revealed a K(m) of 0.43 mM and V(max) of 0.24 micromol/min/mg and a specific activity of 108.9 micromol/min/mg. The isoelectric point of the enzyme was 5.5 by isoelectric focusing and its optimum temperature was 38 degrees C and the enzyme had a maximum activity at approximately pH 8.0. The amino acid composition was also determined for the purified enzyme.
Yang
Hai-ling
HL
College of Life Sciences, Peking University, Beijing 100871, PR China.
Zeng
Qing-yin
QY
Nie
Li-jia
LJ
Zhu
Sheng-geng
SG
Zhou
Xian-wan
XW
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D049872:Bivalvia / Q000201:enzymology*; D005982:Glutathione Transferase / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D007526:Isoelectric Point; D013379:Substrate Specificity
D000596:Amino Acids; D005982:Glutathione Transferase