Partial purification and some biochemical properties of neonatal rat cutaneous glutathione S-transferases.
1. Previous studies have demonstrated the presence of glutathione S-transferases in the skin of rodents and humans. This study represents the first attempt to purify cytosolic glutathione S-transferases from skin of 3-day-old rats. 2. A partial purification of the enzyme was achieved by a two-step procedure: affinity chromatography followed by HPLC. Two peaks, one major (P-1) and one minor (P-2), were resolved by HPLC containing about 82% and 10% of the recovered activity, respectively. 3. The major form exhibited an overall purification of about 2270-fold with a specific activity of about 73 mumoles/min/mg protein towards 1-chloro-2,4-dinitrobenzene. 4. The kinetic data for P-1 yielded mean Km values of 2.39 mM for 1-chloro-2,4-dinitrobenzene and 0.72 mM for reduced glutathione, while the respective average Vmax values were found to be 212 and 101 mumoles/min/mg protein. 5. Significantly inhibition of enzyme activity was noted in the presence of 0.2 mM HgCl2, 0.63 microM 1.2-naphthoquinone, 1.0 microM triphenyltin chloride, and 12.5 microM 17 beta-estradiol-3-sulfate.
Kulkarni
A P
AP
Department of Environmental and Industrial Health, School of Public Health, University of Michigan, Ann Arbor 48109-2029.
Nelson
J L
JL
Radulovic
L L
LL
D000818:Animals; D000831:Animals, Newborn; D002846:Chromatography, Affinity / Q000379:methods; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D005982:Glutathione Transferase / Q000302:isolation & purification* / Q000378:metabolism; D007527:Isoenzymes / Q000302:isolation & purification* / Q000378:metabolism; D007700:Kinetics; D051381:Rats; D011919:Rats, Inbred Strains; D012867:Skin / Q000201:enzymology*
D007527:Isoenzymes; D005982:Glutathione Transferase