A purification scheme is described for the basic and
neutral glutathione S-transferases which occur in human
liver. Three forms with basic isoelectric points,
B-|Bi (pl'8.9), B-|B2 (pi 8.75) and B2B2 (pi 8.4) and two
forms with neutral isoelectric points, N-j (pi 6.1) and
N2 (pi 4.6), were obtained. Not every liver examined
expressed transferase N-] or transferase N2^ An acidic
enzyme from human lung, GST \ (pi 4.8), was included in
the study for the purpose of comparison.
Sodium dodecylsulphate/polyacrylamide-gel electro¬
phoresis demonstrated that each enzyme comprises two subunits
of identical size. As a group the human glutathione
S-transferases are composed of three distinct types of
subunit with different molecular weights. The basic
enzymes (B-]B-j, B1B2 and B2B2) were shown to have a subunit
molecular weight of Mr 25 900, whereas, the neutral enzy¬
mes (N-] and N2) were found to comprise subunits of molecu¬
lar weight Mr 26 500 and the acidic transferase from lung
( X) was shown to be composed of subunits of molecular
weight Mr 24 800. Antisera were raised against each of
these enzymes and they were shown to cross-react only with
enzymes from the same charge/molecular weight group.
Contrary to previous work (Kamisaka e^t_ al_., ( 1975) Eur.
J. Biochem. 60_: 153-161) data are presented to
demonstrate that two distinct basic subunits are
expressed in human liver (B^ and B2). Reversible dissociation and reassociation experiments demonstrated that
transferases B-|Bi and B2B2 could be formed from trans¬
ferase B-1B2. Tryptic-di gest peptide 'maps' showed that
the B-| and B2 subunits possess extensive sequence homo¬
logy, however, seven peptides were recovered from trans¬
ferase B-|Bi that were not recovered from transferase B2B2.
Conversely, four peptides were recovered from transferase
B2B2 that were not recovered from transferase B1B1.
However, all these peptides were recovered from trans¬
ferase B*i B2; this is consistent with the hypothesis that
B1B2 is a hybrid enzyme.
A comparison of the substrate specificities of the
enzymes demonstrated that transferases B -| B1, B-] B2 and
B2B2 had a high peroxidase activity with cumene hydro¬
peroxide, transferases N-| and N2 had a high activity with
trans-4 -phenyl-3 -but en-2- one and transferaseX had a high
activity with ethacrynic acid. The IC50 values obtained
for the basic enzymes demonstrated that the B1 subunit was
much more potently inhibited by tributyltin acetate than
the B2 subunit.
The isoelectric point, molecular weight and high spe¬
cific activity with trans-4-phenyl-3-buten-2-one of trans¬
ferase N-| suggest that it is identical to transferase ]x, a
form that has been previously characterised by Warholm et
al., ( 1983) Biochemistry 22_: 361 0—3617- However, transferanse
N2 is a novel enzyme that has not been described
previously. The properties of this new enzyme indicate
that it also belongs to the neutral group of enzymes.