A glutathione S-transferase (GST) fraction was isolated from cytosol prepar
ed from catfish intestinal mucosa by GSH-agarose affinity chromatography an
d its molecular weight, isoelectric points, substrate specificities and imm
unochemical cross-reactivity were examined. Intestinal GSTs were purified 1
00-fold with respect to cytosolic activity with 1-chloro-2, 4-dinitrobenzen
e and had high activity with ethacrynic acid, (+/-)benzo(a)pyrene-4,5-oxide
, and (+/-)anti-benzo(a)pyrene-7,8-dihydrodiol-9,10-epoxide, but a low acti
vity with 1,2-dichloro-4-nitrobenzene. SDS-polyacrylamide gel electrophores
is revealed the presence of a single band with relative molecular mass of 2
6 700. Gel isoelectric focusing showed a major band with a pI of 8.2. A pol
yclonal antibody prepared against a GST pi-protein isolated from catfish pr
oximal intestine cross-reacted well with the affinity isolated GST fraction
. The catfish antibody also cross-reacted with GST from human placenta whic
h contains predominantly pi-class GST (Mannervik, B., Guthenberg, C., 1981.
Glutathione transferase (human placenta). In: Jakoby, W.B. (Ed.), Methods
in Enzymology, 77. Academic Press, New York, pp. 231-235; Polidoro, G., Dil
lio, C., Arduini, A., Frederici, G., 1981. Molecular and catalytic properti
es of purified glutathione transferase from human placenta. Biochem. Med. 2
2, 247-259; Dao, D.D., Partridge, C.A., Kurosky, A., Awasthi, Y.C., 1982. S
ubunit structure of glutathione-S-transferase of human liver and placenta.
IRSC Med. Sci, Lib. Compend. 10, 175; Dao, D.D., Partridge, C.A., Kurosky,
A., Awasthi, Y.C., 1984. Human glutathione transferase. Characterization of
the anionic forms from lung and placenta. Biochem. J. 221, 33-41), but poo
rly with human liver cytosol. The affinity-isolated protein fraction from w
hole intestine contained proteins that were immunologically related to all
four major classes of human GSTs tested. N-terminal sequence analysis of th
e predominant band obtained by 2D electrophoresis indicated a marked homolo
gy (63-70% identical) to mammalian pi form GST isozymes and very strong sim
ilarity (80%) to a salmon hepatic GST that was designated a pi form (Domine
y, R.J., Nimmo, I.A., Cronshaw, A.D., Hayes, J.D., 1991. The major glutathi
one S-transferase in salmonid fish livers is homologous to the mammalian pi
class GST. Comp. Biochem. Physiol. (B) 100 (1), 93-98). Other bands contai
ned insufficient protein for N-terminal analysis. Taken together, these res
ults indicate that the predominant intestinal GST isoform is related to the
pi-class enzymes, but minor GSTs related to other families are also presen
t. (C) 2000 Elsevier Science B.V. All rights reserved.