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Title
glutathione-dependent bioactivation of xenobiotics.
Author
Dekant W; Vamvakas S
Address
Institut für Toxikologie und Pharmakologie, Universität
Würzburg,
Germany.
Source
Xenobiotica, 23(8):873-87 1993 Aug
Abstract
glutathione conjugation has been identified as an important
detoxication reaction. However, in recent years several
glutathione-dependent bioactivation reactions have been identified.
Current knowledge on the mechanisms and the possible biological
importance of these reactions are discussed. 1. Dichloromethane
is
metabolized by glutathione conjugation to formaldehyde via
S-(chloromethyl)glutathione. Both compounds are reactive intermediates
and may be responsible for the dichloromethane-induced tumorigenesis
in
sensitive species. 2. Vicinal dihaloalkanes are transformed by
glutathione S-transferase-catalyzed reactions to mutagenic and
nephrotoxic S-(2-haloethyl)glutathione S-conjugates. Electrophilic
episulphonium ions are the ultimate reactive intermediates formed.
3.
Several polychlorinated alkenes are bioactivated in a complex,
glutathione-dependent pathway. The first step is hepatic glutathione
S-conjugate formation followed by cleavage to the corresponding
cysteine S-conjugates, and, after translocation to the kidney,
metabolism by renal cysteine conjugate beta-lyase. Beta-Lyase-dependent
metabolism of halovinyl cysteine S-conjugates yields electrophilic
thioketenes, whose covalent binding to cellular macromolecules
is
responsible for the observed toxicity of the parent compounds.
4.
Finally, hepatic glutathione conjugate formation with hydroquinones
and
aminophenols yields conjugates that are directed to
gamma-glutamyltransferase-rich tissues, such as the kidney, where
they
undergo alkylation or redox cycling reactions, or both, that
cause
organ-selective damage.
Title
Significance of an unusually low Km for glutathione in glutathione
transferases of the alpha, mu and pi classes.
Author
Meyer DJ
Address
Department of Biochemistry and Molecular Biology, University
College
London, UK.
Source
Xenobiotica, 23(8):823-34 1993 Aug
Abstract
1. Interactions of glutathione transferases (GST) of the alpha,
mu and
pi classes with glutathione (GSH) and glutathione conjugates
(GS-X) are
in contrast with those of a GST of the theta class (GST5-5).
2. GST 5-5
has a Km for GSH of approx. 5 mM. Thus Km/ambient [GSH] is approx.
1,
within the range of Km/ambient [s] of glycolytic enzymes. GSTs
of the
alpha, mu and pi classes yield much lower values of Km for GSH
(approx.
0.1 mM) hence Km/ambient [s] is significantly lower than those
of most
(non-GST) enzymes (p < 0.025). 3. GSTs of the alpha, mu and
pi classes
are sensitive to inhibition by GS-X (i.e. product) and GS-X analogues.
GST 5-5 is not. 4. Rate enhancements up to 10(10), similar to
an
average enzyme (10(8)-10(12)), are seen in catalysis by GST 5-5,
but
not in catalysis by GSTs of alpha, mu and pi classes (> 10(7)).
5.
Comparisons of primary structure indicate that theta class GSTs
may
have a decreased binding of the glu-alpha-amino- and gly-COO(-)-groups
of GSH compared with GSTs of the other classes. 6. It is concluded
that
GSTs of alpha, mu and pi classes have evolved towards increased
product
binding at the expense of catalytic efficiency. Thus GSH is uniquely
utilized both as a nucleophile and a 'tag' which can be used
to bind
and sequester product particularly during GSH-depletion. This
interpretation unifies the catalytic and binding properties of
these
GSTs and alters their perceived role in detoxication.
Title
Deficient glutathione peroxidase activity in preeclampsia is
associated
with increased placental production of thromboxane and lipid
peroxides.
Author
Walsh SW; Wang Y
Address
Department of Obstetrics and Gynecology, Medical College of Virginia,
Virginia Commonwealth University, Richmond 23298-0034.
Source
Am J Obstet Gynecol, 169(6):1456-61 1993 Dec
Abstract
OBJECTIVE: Thromboxane and lipid peroxide levels are abnormally
increased in preeclamptic placentas. The cause of this increase
is not
known. Peroxides stimulate prostaglandin H2 synthase to increase
thromboxane and oxygen radicals, which increase lipid peroxides.
glutathione peroxidase inactivates peroxides, thereby decreasing
peroxide stimulation of prostaglandin H synthase. If glutathione
peroxidase activity were deficient, then peroxides could increase,
leading to increased stimulation of prostaglandin H synthase,
resulting
in increased production of thromboxane and lipid peroxides. The
following study tested this hypothesis. STUDY DESIGN: Placental
tissues
from 11 normal and 11 preeclamptic women were immediately frozen
in
liquid nitrogen after delivery. One gram of tissue from each
placenta
was homogenized for analysis. Placental tissues were also obtained
from
six normal pregnancies for incubation with a glutathione peroxidase
inhibitor, N-ethylmaleimide. Samples were analyzed for glutathione
peroxidase activity, lipid peroxides by hydrogen peroxide equivalents,
thromboxane by thromboxane B2, and prostacyclin by 6-keto-prostaglandin
F1 alpha. RESULTS: glutathione peroxidase activity was significantly
lower in preeclamptic than in normal placentas (9.41 +/- 0.05
vs 13.41
+/- 0.63 units/gm, p < 0.001, mean +/- SE). Lipid peroxides
and
thromboxane were significantly higher in preeclamptic than in
normal
placentas (hydrogen peroxide equivalents 4.23 +/- 0.32 vs 2.84
+/- 0.27
mumol/gm, p < 0.01; thromboxane B2 215 +/- 31 vs 138 +/- 15
ng/gm, p <
0.05), whereas prostacyclin was significantly lower
(6-keto-prostaglandin F1 alpha 23.3 +/- 2.2 vs 41.6 +/- 6.0 ng/gm,
p <
0.01). Inhibition of glutathione peroxidase activity in normal
placentas resulted in a dose-response increase in placental production
of both lipid peroxides and thromboxane without affecting prostacyclin,
so the ratio of thromboxane to prostacyclin increased progressively
with inhibition of glutathione peroxidase. CONCLUSIONS: glutathione
peroxidase activity is significantly lower, and lipid peroxides
and
thromboxane are significantly higher, in preeclamptic placentas
compared with normal placentas. Inhibition of glutathione peroxidase
activity in normal placentas results in significantly increased
production of lipid peroxides and thromboxane and an increase
in the
thromboxane to prostacyclin ratio. We speculate that in normal
placentas, glutathione peroxidase limits prostaglandin H synthase
activity by reducing the amount of peroxide present, thus reducing
peroxide stimulation of prostaglandin H synthase. In preeclampsia
deficient glutathione peroxidase activity results in in increased
peroxide levels leading to increased stimulation of prostaglandin
H
synthase, which results in increased production of lipid peroxides
and
thromboxane.
Title
The subcellular distribution of the glutathione system enzymes
in the
brain tissue of the rat
Author
Kozhemiakin LA; Bulavin DV; Udintsev AV; Smirnov VV
Source
Tsitologiia, 35(6-7):58-63 1993
Abstract
Using the percoll density gradient method for rapid isolation
of
organelles the distribution of glutathione system enzymes in
the rat
brain was characterized including glutathione S-transferase (GST),
glutathione peroxidase (GPx), and glutathione reductase (GR).
In the
fraction of mitochondria, the shares of GPx, GR, and GST were
found to
be as much as 92, 61, and 15% from their respective general activities.
No enzymatic activity was found in lysosomes. Fractions of organelles
with low density (microsomes, Golgi complex, cytoplasmic membranes)
were cross-contaminated. Nevertheless, on the base of the established
dynamics of distribution of the marker enzyme activity among
various
cell organelles it is suggested that, besides mitochondria, the
glutathione system enzymes may be localized primarily in the
endoplasmic reticulum, rather than in other organelles.
Title
Correction of glutathione deficiency in the lower respiratory
tract of
HIV seropositive individuals by glutathione aerosol treatment.
Author
Holroyd KJ; Buhl R; Borok Z; Roum JH; Bokser AD; Grimes GJ; Czerski
D;
Cantin AM; Crystal RG
Address
Pulmonary Branch, National Heart, Lung, and Blood Institute,
National
Institutes of Health, Bethesda, Maryland 20892.
Source
Thorax, 48(10):985-9 1993 Oct
Abstract
BACKGROUND--Concentrations of glutathione, a ubiquitous tripeptide
with
immune enhancing and antioxidant properties, are decreased in
the blood
and lung epithelial lining fluid of human immunodeficiency virus
(HIV)
seropositive individuals. Since the lung is the most common site
of
infection in those who progress to AIDS it is rational to consider
whether it is possible to safely augment glutathione levels in
the
epithelial lining fluid of HIV seropositive individuals, thus
potentially improving local host defence. METHODS--Purified reduced
glutathione was delivered by aerosol to HIV seropositive individuals
(n
= 14) and the glutathione levels in lung epithelial lining fluid
were
compared before and at one, two, and three hours after aerosol
administration. RESULTS--Before treatment total glutathione
concentrations in the epithelial lining fluid were approximately
60% of
controls. After three days of twice daily doses each of 600 mg
reduced
glutathione, total glutathione levels in the epithelial lining
fluid
increased and remained in the normal range for at least three
hours
after treatment. Strikingly, even though > 95% of the glutathione
in
the aerosol was in its reduced form, the percentage of oxidised
glutathione in epithelial lining fluid increased from 5% before
treatment to about 40% three hours after treatment, probably
reflecting
the use of glutathione as an antioxidant in vivo. No adverse
effects
were observed. CONCLUSIONS--It is feasible and safe to use aerosolised
reduced glutathione to augment the deficient glutathione levels
of the
lower respiratory tract of HIV seropositive individuals. It is
rational
to evaluate further the efficacy of this tripeptide in improving
host
defence in HIV seropositive individuals.
Title
Inhibition of oral carcinogenesis by glutathione.
Author
Trickler D; Shklar G; Schwartz J
Address
Department of Oral Medicine and Oral Pathology, Harvard School
of
Dental Medicine, Boston, MA 02115.
Source
Nutr Cancer, 20(2):139-44 1993
Abstract
Forty young adult Syrian hamsters (Mesocricetus auratus) were
divided
into four groups of 10 animals each. In Group 1 (tumor control),
the
right buccal pouches were painted three times per week with a
0.5%
solution of 7,12-dimethylbenz[a]anthracene (DMBA) in heavy mineral
oil
(USP) with a no. 4 sable brush. In Group 2 (experimental group),
the
right buccal pouches were painted with DMBA, as in Group 1. In
addition, Group 2 received 1 mg of reduced glutathione in 0.5
ml of
mineral oil three times per week on days alternate to the DMBA
application. The glutathione was administered systemically by
mouth
with a pipette. Group 3 received only glutathione, and Group
4 was
untreated (control groups). Animals were sacrificed after 14
weeks, and
tumors were counted and measured. Both right and left pouches
were
photographed, excised, fixed in formalin, sectioned in paraffin,
and
studied histologically. The animals receiving glutathione demonstrated
significantly fewer and smaller tumors. The mean tumor burden
was 315
mm3 in the glutathione-treated group and 3,040 mm3 in the untreated
group. The statistical significance by Student's t test was <
or =
0.0001. Histological study also revealed significantly fewer
areas of
dysplastic leukoplakia in the group treated with glutathione.
This
study represents the first demonstration of the anticancer effect
of
systemically administered reduced glutathione.
Title
Maintenance of neuronal glutathione by glial cells.
Author
Sagara JI; Miura K; Bannai S
Address
Department of Biochemistry, University of Tsukuba Medical School,
Ibaraki, Japan.
Source
J Neurochem, 61(5):1672-6 1993 Nov
Abstract
glutathione levels in neurons and glial cells were investigated
in a
neuronal-glial coculture and in separate cultures. Brain cell
suspensions obtained from cerebral hemispheres of fetal rats
were
cultured, and after 5 days the glutathione content of this cell
population, consisting mainly of neurons and astroglial cells,
was 23.0
nmol/mg of cell protein, with a significantly high content in
glial
cells (28.0 nmol/mg of protein) in comparison with neurons (18.8
nmol/mg of protein). When the neurons and glial cells were separated
and recultured in fresh medium, neuronal glutathione rapidly
decreased,
whereas glial glutathione remained unchanged. cysteine is a
rate-limiting precursor for glutathione synthesis, and its level
was
also decreased in neurons, but not in glial cells. cysteine was
taken
up rapidly by both neurons and glial cells, but cystine was taken
up
only by glial cells. This accounts for the rapid decrease of
glutathione in the cultured neurons, because the culture medium
contains cystine, but not cysteine. It was also found that the
cultured
glial cells released cysteine into the medium. These results
suggest
that neurons maintain their glutathione level by taking up cysteine
provided by glial cells.
Title
Site-directed mutagenesis of glutathione S-transferase YaYa.
Mapping
the glutathione-binding site.
Author
Wang RW; Newton DJ; Johnson AR; Pickett CB; Lu AY
Address
Department of Drug Metabolism, Merck Research Laboratories, Rahway,
New
Jersey 07065.
Source
J Biol Chem, 268(32):23981-5 1993 Nov 15
Abstract
Previous studies from our laboratory have shown that aspartic
acid 101
plays an important role in glutathione interaction to rat glutathione
S-transferase YaYa, while tyrosine 9 is directly involved in
catalysis.
Based on the available structural information, site-directed
mutagenesis was conducted to examine the function of arginine,
lysine,
glutamine, and proline residues surrounding the GSH binding pocket.
Arginine mutants R13K, R15K, R20K, and R20I retained partial
enzymatic
activities, while R13I and R15I lost most of their activities.
Kinetic
studies showed a marked increase in Km toward GSH for R15I suggesting
that arginine 15 contributes significantly to the binding of
GSH in the
active site of glutathione S-transferase YaYa. A drastic decrease
in
enzymatic activities for R13I suggested the importance of the
charged
group of arginine 13 either in maintaining the structural integrity
of
the enzyme or in serving a vital role in enzymatic function.
Replacement of glutamine 54 and 67 with glutamic acid or asparagine
resulted in decreased enzymatic activities. Moreover, an 11-,
17-, and
9-fold increase in Km values toward GSH for mutant Q54E, Q54N,
and Q67N
was observed, respectively. These results suggested that glutamine
54
and 67 also contributed significantly to the binding of GSH.
Proline at
position 56 appears to be important for maintaining the structural
integrity of the enzyme since mutants P56A and P56F were much
less
active and extremely less stable than that of the wild type enzyme.
Both lysine mutants, K45R and K45I, exhibited substantially higher
catalytic efficiencies toward both 1-chloro-2,4-dinitrobenzene
and GSH
than the wild type enzyme. Our data clearly show that lysine
45 is not
an essential residue for catalysis nor for GSH binding in glutathione
S-transferase YaYa. |
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