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I Listowsky

Publications and source records attributed to I Listowsky.

At least 37 records · Page 2Linked to original sources
PubMed1989

Glutathione-S-transferases are major cytosolic thyroid hormone binding proteins.

Thyroid hormone binding proteins of rat liver cytosol were characterized. Glutathione-S-transferases were identified among major cytosolic proteins adsorbed by thyroxine affinity matrices. The Ya and Yb subunits of the glutathione-S-transferases were also principal proteins of cytosol covalently labeled with 3,3',5-triiodo-L-thyronine (T3) or 3,3',5,5'-tetraiodo-L-thyronine (T4) by photoaffinity methods. T3 and T4, but not L-thyronine or iodinated tyrosines, were bound with high affinity to purified glutathione-S-transferases and were potent inhibitors of their enzymatic activities. These results suggest that glutathione-S-transferases have the potential to function in the intracellular binding and transport of thyroid hormones. The proteins provide a means for regulating the action and metabolism of thyroid hormones by acting as high capacity binding components.

Affinity Labels↗
PubMed1989

Differential localization of glutathione-S-transferase Yp and Yb subunits in oligodendrocytes and astrocytes of rat brain.

Glutathione-S-transferase Yb subunits were recently identified in rat brain and localized to astrocytes, ependymal cells lining the ventricles, subventricular zone cells, and tanycytes. Another isoform, Yp (pi family), was detected in rat brain by immunoblotting, and its mRNA was detected by Northern hybridizations. Double immunofluorescence localized Yb and Yp in different glial cells. The strongly Yp-positive cells were identified as oligodendrocytes by virtue of their arrangement in rows in white-matter tracts, colocalization in strongly carbonic anhydrase-positive cells, and association with myelinated tracts in the corpus striatum. Ependymal cells in the choroid plexus and ventricular lining were also strongly Yp positive, whereas Yb was not detected in the choroid plexus. The occurrence of Yp at low levels in astrocytes was indicated after immunostaining by a sensitive peroxidase-antiperoxidase method, which revealed weak staining of those cells in the molecular layer of the cortex. The data suggest that Yb and Yp subunits are primarily localized to astrocytes and oligodendrocytes, respectively, and that both are absent from neurons. The glutathione-S-transferase in oligodendrocytes may participate in the removal of toxins from the vicinity of the myelin sheath. The finding of glutathione-S-transferases in ependymal cells and astrocytes in the brain also suggests that this enzyme could be a first line of defense against toxic substances.

Animals↗
PubMed1988

Expression of an enzymatically active Yb3 glutathione S-transferase in Escherichia coli and identification of its natural form in rat brain.

Glutathione S-transferases containing Yb3 subunits are relatively uncommon forms that are expressed in a tissue-specific manner and have not been identified unequivocally or characterized. A cDNA clone containing the entire coding sequence of Yb3 glutathione S-transferase mRNA was incorporated into a pIN-III expression vector used to transform Escherichia coli. A fusion Yb3-protein containing 14 additional amino acid residues at its N terminus was purified to homogeneity. Recombinant Yb3 was enzymatically active with both 1-chloro-2,4-dinitrobenzene and 1,2-dichloro-4-nitrobenzene as substrates but lacked glutathione peroxidase activity. Substrate specificity patterns of recombinant Yb3 were more limited than those of glutathione S-transferase isoenzymes containing Yb1- or Yb2-type subunits. Peptides corresponding to unique amino acid sequences of Yb3 as well as a peptide from a region of homology with Yb1 and Yb2 subunits were synthesized. These synthetic peptides were used to raise antibodies specific to Yb3 and others that cross-reacted with all Yb forms. Immunoblotting was utilized to identify the natural counterpart of recombinant Yb3 among rat glutathione transferases. Brain and testis glutathione S-transferases were rich in Yb3 subunits, but very little was found in liver or kidney. Physical properties, substrate specificities, and binding patterns of the recombinant protein paralleled properties of the natural isoenzyme isolated from brain.

Amino Acid Sequence↗
PubMed1988

Characterization and localization of glutathione-S-transferases in rat brain and binding of hormones, neurotransmitters, and drugs.

Rat brain glutathione-S-transferases are rich in Yb type subunits with major RNA transcripts coding for a relatively uncommon Yb3 form. The Yb-containing isoenzymes of brain cytosol bind glucocorticoids and are covalently labeled with dexamethasone 21-methanesulfonate. Certain neurotransmitters, hormones, and drugs, such as serotonin, dopamine, glucocorticoids, thyroxine, apomorphine, and benzodiazepine derivatives, are effective inhibitors of brain glutathione transferase activity. Immunocytochemical studies show that Yb forms are localized in ependymal cells, subventricular zone cells, astrocytes, tanycytes, and astrocyte foot processes on blood vessels, but Yb was not detected in oligodendrocytes or neurons. Based on their localization and binding properties, brain glutathione-S-transferases have the potential to function in intracellular binding of a variety of compounds and thereby govern their uptake and release in brain, transport to neurons, as well as in their detoxification.

Animals↗
PubMed1988

Developmental regulation of glutathione S-transferases.

1. cDNA probes for individual isoenzymes of rat glutathione S-transferases were used to determine steady state levels of their mRNAs in liver and brain during development. 2. Foetal livers were enriched in Yp transcripts (that are characteristic of hyperplastic nodules and hepatocellular carcinomas), but these forms decreased after the first week of postnatal development and were not detected in adult livers. In contrast, in adult brains Yp levels increased. 3. Ya forms that were present at low levels in foetal and neonatal livers, increased markedly during development. Ya was not detected in brain. 4. The Yb1 and Yb2 GSTs were present in higher amounts than Ya in foetal livers, and these isoenzymes also increased in adults.

Age Factors↗
PubMed1987

Selective expression of a unique glutathione S-transferase Yb3 gene in rat brain.

A cDNA clone from a rat brain lambda gt11 expression vector library contained the entire open reading frame, the 3' untranslated sequence, and 18 nucleotides of the 5' untranslated region of a Yb-type glutathione S-transferase that was clearly distinct from previously characterized liver forms. This form was designated as Yb3. Primer extension analysis indicated that the mRNA for Yb3 extends 118 nucleotides upstream of the translation initiation codon, and its total length of 1308 nucleotides is substantially larger than the transcripts of any of the liver glutathione S-transferases described thus far. The open reading frame of 654 nucleotides of Yb3 is the same as that of liver Yb1 and Yb2, with greater than 80% sequence homology. The amino acid sequence derived for Yb3 had regions conserved in all forms of Yb glutathione S-transferase, some other amino acid residues common to either Yb1 or Yb2, and 28 substitutions unique to Yb3. The 533 nucleotides in the 3' untranslated region of Yb3 were longer and not homologous to the 3' noncoding ends of the previously described glutathione S-transferases. A second clone from the brain library was virtually identical to that of the gene encoding the liver Yb1 form of the enzyme. Northern blot analyses of rat brain poly(A)+ RNA with specific Yb1 and Yb3 probes showed that the Yb3 form hybridized with a 1300-nucleotide mRNA, Yb1 with a separate 1100-nucleotide component, and that the two probes did not cross-hybridize. Moreover, while Yb3 is a minor component in liver, testis, and heart with even lesser amounts in spleen, lung, and kidney, it is a major glutathione S-transferase transcript in rat brain.

Amino Acid Sequence↗
PubMed1986

A subclass of glutathione S-transferases as intracellular high-capacity and high-affinity steroid-binding proteins.

The distribution of glucocorticoids incubated with rat liver cytosol preparations or administered in vivo to adrenalectomized rats was analysed by chromatographic procedures. Corticosterone or dexamethasone was co-eluted with Yb-type GSH S-transferases in anion-exchange and gel-permeation chromatography systems, and these glucocorticoids also were bound to Yb forms in analyses by immunoadsorbent and lysyl-GSH affinity matrices. Pretreatment of cytosol with lysyl-GSH to extract GSH S-transferases or incubation with excess bilirubin, which is expected to compete with steroids for binding to the protein, yielded preparations that were devoid of this major steroid-binding component. In mixtures of the multiple rat GSH S-transferases, corticosterone preferentially interacted with Yb forms rather than Ya and Yc subgroups. All of the multiple Yb forms resolved by chromatofocusing procedures retained the steroid-binding capacity. It is suggested that these abundant proteins can account for a considerable share of intracellular glucocorticoid binding and represent a high-affinity non-saturable binding component with potential to function in steroid-hormone metabolism and action.

Animals↗
PubMed1985

Temperature dependent redistribution among the multiple forms of rat Yb-glutathione-S-transferase.

Anionic (Yb) rat liver glutathione-S-transferases are susceptible to temperature or pH dependent transitions to more basic forms of this class of proteins. At elevated temperatures (25-30 degrees) or at pH values above 9.0 the protein is rapidly and irreversibly converted to forms that are no longer retained by anion exchange resins and display basic components in chromatofocusing systems, because bound glutathione is removed at the higher temperatures or pH. Sharp increases in enzymatic activity with 1,2-dichloro-4-nitrobenzene as a substrate, accompany the temperature induced changes. Microheterogeneity patterns for this protein are contingent upon these interconversions, and the results explain apparent variations in relative amounts of the multiple forms under different conditions in terms of glutathione binding.

Animals↗
PubMed1985

Concentration-dependent sedimentation properties of ferritin: implications for estimation of iron contents of serum ferritins.

Serum ferritins from various sources sedimented at lower densities than tissue ferritins in sucrose gradient centrifugation systems. The sedimentation patterns of ferritins, however, were shown to be dependent on the concentration of the protein; as the concentration decreased the protein appeared to sediment at lower densities. Thus, at the low concentration levels usually used for analysis of serum ferritin, tissue ferritins also sedimented in the same lower density regions. Iron labeling experiments indicated that the sedimentation changes upon dilution were not due to release of iron or was there any indication that the protein dissociated into subunits. The anomalous sedimentation behavior of serum ferritin should therefore not be interpreted in terms of its iron content. The disclosure that serum ferritins may have full complements of iron is counter to the prevalent view that serum ferritins are low iron forms and has potential implications with regard to the sources and possible function of this protein in the circulation.

Animals↗
PubMed1985

Identification of Yb-glutathione-S-transferase as a major rat liver protein labeled with dexamethasone 21-methanesulfonate.

Dexamethasone 21-methanesulfonate, an affinity label for glucocorticoid-binding proteins, was incubated with rat liver cytosol preparations. The predominant covalently labeled component was identified as Yb-glutathione-S-transferase on the basis of chromatographic properties, electrophoretic mobility, and specific retention by an anti-Yb-immunoadsorbent. Affinity labeling of this protein was blocked by excess dexamethasone. Preferential reactivity of dexamethasone 21-methanesulfonate with the Yb subclass of glutathione-S-transferase (glutathione transferase, EC 2.5.1.18) was also evident with mixtures containing the multiple forms of the enzyme. Yb-glutathione-S-transferase, the nonsaturable glucocorticoid-binding component of rat liver cytosol should, therefore, be reclassified; because of its high concentration and selective interaction with steroids, this enzyme may be an intracellular glucocorticoid-binding protein and, thereby, influence transport, metabolism, and action of the steroids.

Affinity Labels↗
PubMed1984

Preferential binding of steroids by anionic forms of rat glutathione S-transferase.

Rat liver glutathione S-transferases with isoelectric points near 6.7 were resolved from more basic forms of the protein. This anionic fraction represented about 30% of the total activity in liver with 1-chloro-2,4-dinitrobenzene and was the preponderant form utilizing trans-4-phenyl-3-butene-2-one as a substrate. The anionic transferases are dimeric proteins composed of two subunits designated as Yb and were distinguished from the cationic transferases on the basis of structural, immunological, and binding properties. Amino acid compositions and immunological properties of the anionic protein were similar to those of glutathione S-transferases A and C. The anionic forms had substantially less ordered secondary structure than cationic forms composed of subunits Ya and Yc. Stoichiometric ratios of two high affinity binding sites per dimer, also differentiated between the anionic and all of the cationic transferases which bind only a single mole of ligand. Affinity matrices composed of corticosterone or cholate, and circular dichroism methods, were used to demonstrate selective binding of steroids and bile acids to the anionic glutathione S-transferases. Glucocorticoids and progestins were shown to bind with high affinity whereas estrogens were bound at distinct lower affinity sites. In contrast to the cationic transferases, glutathione had no effect on binding of the steroids to the anionic forms, which suggested that these proteins have the capacity to bind these substances even in a milieu with high concentrations of glutathione.

Amino Acids↗
PubMed1984

Distinctions between the multiple cationic forms of rat liver glutathione S-transferase.

Three cationic glutathione S-transferase forms isolated from rat liver were characterized as dimers that originated from different combinations of two subunit types, Ya and Yc. The cationic forms were purified using lysyl glutathione affinity matrices and were chromatographically resolved from anionic glutathione S-transferases that contain Yb subunits. The three classes of cationic transferase exhibited similar specific activities with 1-chloro-2,4-dinitrobenzene as a substrate, all forms cross-reacted with antibodies to glutathione S-transferase B, and all had comparable secondary structures and tryptophan fluorescence properties. In spite of those similarities, the Yc-containing forms were clearly distinguishable from Ya forms on the basis of characteristic differences in circular dichroic patterns associated with their aromatic side chains. All cationic transferases bound bilirubin with stoichiometric ratios of 1 mol/dimeric protein molecule, but discrete differences in mode of binding were ascribed to forms containing Ya subunits as compared to Yc dimers. Binding to Yc forms was of lower affinity and may be associated with the catalytic region of the protein since glutathione effectively displaced bilirubin from the Yc component.

Animals↗
PubMed1983

Iron uptake and regulation of ferritin synthesis by hepatoma cells in hormone-supplemented serum-free media.

Iron, as ferric nitrilotriacetate or ferric ammonium citrate, was administered to rat hepatoma cells (H4AZC2) that were grown in serum-containing media or in hormone-supplemented defined media on collagen matrices. High levels of iron either retarded growth or were cytotoxic, so conditions were established for maximum iron loading where cells survived at near normal growth rates. In all cases, cells exposed to iron produced more ferritin than those grown in its absence, and elevated ferritin levels were paralleled by higher intracellular iron contents. Cells grown in serum-free media, however, took up iron more rapidly than corresponding cells in serum-supplemented media, and intracellular iron and ferritin also reached much higher levels. During exponential growth stages, for example, ferritin levels in iron-stimulated cells were 35-fold greater than those in control cells. These results indicate that transferrin is not required as an iron donor as the inorganic iron was taken up effectively and utilized to stimulate ferritin synthesis. Twenty-four hours after iron administration, endocytotic mechanisms were evident by the appearance of coated vesicles and pits and visible cytoskeletal structures. Subsequently, clusters of iron micelles appeared in the cell. Ferritin isolated from iron-overloaded cells were rich in L-type subunits, but newly synthesized ferritins in iron-stimulated or control cells had almost equivalent amounts of heavy and light subunit constituents.

Animals↗
PubMed1981

Structure, assembly, conformation, and immunological properties of the two subunit classes of ferritin.

The two subunit types of human liver ferritin were purified to homogeneity. Both subunits reassembled in a well-defined manner and formed spherical particles that resembled natural apoferritin in electron micrographs. Affinity chromatography methods were employed to obtain preparations of antibodies that interacted exclusively either with the H or with the L polypeptides, demonstrating that distinct immunological properties may be ascribed to each subunit of ferritin. The amino acid compositions of the subunits were similar, but the larger H subunit had fewer leucine, phenylalanine, and arginine residues. It is therefore improbable that H subunits undergo proteolytic processing and are precursors for L subunits. Circular dichroism data indicated that homopolymers assembled from L-type subunits had substantially more ordered secondary structures and greater alpha-helical contents than their H counterparts. Small differences in the environment of tryptophan residues were evident from fluorescence spectra of each homopolymer. In isoelectric focusing experiments reassembled H or L homopolymers migrated as families of proteins within discrete pI ranges which are probably representative of subpopulations of each subunit type. The H homopolymer focused at lower pI's than the L component. These data substantiate the contention that both subunits are authentic polypeptide moieties of ferritin with some common structural features, but the results also underscore prominent dissimilarities in their properties.

Apoferritins↗
PubMed1980

Assembly of intra- and interspecies hybrid apoferritins.

An intraspecies hybrid apoferritin was assembled by mixing subunits of horse heart ferritin, which consists mainly of H-type subunits, and horse spleen ferritin, in which L-type subunits predominate. Interspecies hybrid apoferritins were reconstituted from subunits of human liver-horse spleen ferritins and from rat liver-horse spleen ferritins. All the hybrid ferritins migrated as single zones with electrophoretic mobilities intermediate between those of the parent ferritins. Isoelectric focusing data and immunological patterns were consistent with the view that the reassembled apoferritins were composite molecules that contained subunits from each of the interacting forms. Reconstitution occurred in a random manner, as there was no apparent preference for assembly of homologous subunits. These results suggest that intersubunit interaction domains and recognition mechanisms that dictate formation of the highly specific quaternary structure assumed by this protein are common for different species of ferritins.

Animals↗
PubMed1980

Ligandin: an adventure in liverland.

Ligandin is an abundant soluble protein which has a t 1/2 of 2--3 days, is induced by many drugs and chemicals, and is stabilized in the absence of thyroid hormone. The protein is strategically concentrated in cells associated with transport and detoxification of many endogenous ligands, such as bilirubin, and exogenous ligands, such as drugs and chemicals. The protein is a dimer in rat liver. Whether the dimer is a primary gene product or at least two genes are involved is not known. The protein has broad, low affinity catalytic activity as a GSH-S-transferase for many ligands having electrophilic groups and hydrophobic domains. It catalyzes formation of GSH conjugates, non-covalently binds some ligands prior to their biotransformation or excretion in bile, and covalently binds other ligands, such as activated carcinogens. Recent studies include the possible role of ligandin in chemical carcinogenesis, diagnosis of inflammatory and neoplastic disease of the liver and kidney, and participation in intracellular transport. Although some of the roles that have been outlined are speculative, any single function is important. The GSH-S-transferases are primitive enzymes and non-specific binding proteins but "it is precisely their simplistic design that allows such protean serviceability". Ligandin illustrates a group of hepatic disposal mechanisms which involve bulk transport of ligands. Although specific uptake and transport mechanisms have been described for several hormones which enter the hepatocyte in small quantities and regulate intermediary metabolism and, possibly, cell maturation, bulk transport of ligands into, through and out of the liver involves mechanisms which accomodate many metabolites, drugs and chemicals of diverse structure. The liver is bathed in sewage which contains what we ingest or are injected with and potentially toxic products of intestinal microorganisms. The chemical formulas of the many substances which are metabolized by the liver provide a horror show of potentially reactive and toxic metabolites, mutagens and carcinogens. Despite this alimentary "Love Canal", we and our livers do remarkably well. These hepatic disposal mechanisms, as exemplified by ligandin, evolved in ancient times. They are present, albeit sluggishly, in insects and ancient elasmobranchs. Hepatic uptake and removal mechanisms of high capacity, modest affinity and broad substrate range permit us to live in what has probably always been a threatening world.

Animals↗