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G H Tait

Publications and source records attributed to G H Tait.

At least 19 recordsLinked to original sources
PubMed1991

17 Beta-hydroxysteroid dehydrogenases in human breast tissues: purification and characterization of soluble enzymes and the distribution of particulate and soluble forms in adipose, non-adipose and tumour tissues.

17 Beta-Hydroxysteroid dehydrogenase (17 beta-HSD) is present in multiple forms in human breast tissue. One soluble form, with a molecular weight of approximately 35 kDa, was purified to near homogeneity from whole normal breast tissue. This form catalysed the oxidation of oestradiol and the reduction of oestrone, with NADP+ and NADPH as the preferred coenzymes. Three other soluble forms with higher molecular weights (in the range 50-80 kDa) were isolated. They catalysed the oxidation of oestradiol but not the reduction of oestrone, and all of them had properties very different from those of the low molecular weight enzyme. Activities of 17 beta-HSD were measured in particulate and soluble fractions from normal breast adipose and non-adipose tissues, and from breast tumours obtained from post-menopausal women, in the oxidative direction with NAD+ and NADP+ as coenzymes and in the reductive direction with NADH and NADPH as coenzymes. Particulate fractions from tumours had much higher oxidative and reductive activities than those from normal tissues. Soluble fractions from tumours had higher oxidative activities than those from the normal tissues but similar reductive activities. The major soluble form of 17 beta-HSD in adipose tissue was the 35 kDa enzyme which had both oxidative and reductive activities. In contrast, the majority of the soluble activity in non-adipose tissue was due to enzymes, with molecular weights in the range 50-80 kDa, which had oxidative activity only. The soluble fractions of tumours, like those of non-adipose tissue, contained enzymes with molecular weights in the range 50-80 kDa. In addition, they contained a 35 kDa enzyme with properties different from those of the enzyme with the same molecular weight present in adipose tissue.

17-Hydroxysteroid Dehydrogenases↗
PubMed1989

Multiple forms of 17 beta-hydroxysteroid oxidoreductase in human breast tissue.

17 beta-Hydroxysteroid oxidoreductase, the enzyme that catalyses the interconversion of oestradiol and oestrone, is known to be present in human breast tissue. However, it is not known whether one or more forms of the enzyme is present. Homogenates of breast adipose tissue and breast glandular tissue were fractionated and fractions assayed in the oxidative direction with NAD+ and NADP+ as coenzymes, and in the reductive direction with NADH and NADPH as coenzymes. Ultracentrifugation of homogenates showed that there was membrane-bound activity and soluble activity. The soluble activity was due to a number of forms of the enzyme with different molecular weights, three in breast adipose tissue and two in breast glandular tissue, as shown by fractionation with (NH4)2SO4 followed by chromatography on Sephadex G-200. The forms of the enzyme isolated differed in their affinities for substrates and coenzymes and in the relative rates at which they catalysed the oxidative and reductive reactions. Preliminary experiments with breast tumours showed that they also contained membrane-bound activity and more than one soluble form of the enzyme.

17-Hydroxysteroid Dehydrogenases↗
PubMed1988

Paracrine influence of human breast stromal fibroblasts on breast epithelial cells: secretion of a polypeptide which stimulates reductive 17 beta-oestradiol dehydrogenase activity.

We have previously shown that a stroma-associated paracrine influence may occur in the human breast. In particular, human breast fibroblasts secrete a factor which stimulates reductive 17 beta-oestradiol dehydrogenase (HSD) activity, thereby regulating tissue concentrations of 17 beta-oestradiol. We report here the results of experiments designed to establish the nature of the enzyme activity stimulating factor. In vitro cell culture techniques were used, in which human breast fibroblast-conditioned medium was used to grow the human breast cancer cell line, MCF-7, for 6 days, after which the reductive HSD activity of the monolayers was assessed. The fibroblastic reductive HSD stimulating factor was found to be a trypsin-sensitive polypeptide. The polypeptide eluted from a Sephadex G-75 column as a peak corresponding to a molecular weight of about 50 kDa. The polypeptide exerts its effects by altering the Vmax of 2 of the cytosolic forms of HSD within MCF-7 cells. This is achieved by a protein-synthesis-dependent but calmodulin-independent mechanism. These results provide further evidence of a paracrine effect by stromal tissue within the human breast and have important implications with respect to the aetiology and treatment of breast cancer.

17-Hydroxysteroid Dehydrogenases↗
PubMed1981

Species variations in the renal and hepatic conjugation of 3-phenoxybenzoic acid with glycine.

1. 3-Phenoxy[14C]benzoyl-CoA has been chemically synthesized, purified and characterized by field-desorption mass spectrometry. Biological activity of the purified thioester was greater than 92%. 2. The two enzymic steps involved in the conjugation of 3-phenoxybenzoic acid (3PBA) with glycine have been investigated in hepatic and renal tissues from various mammalian species. 3. A 10- to 300-fold excess of acyl-CoA: glycine N-acyltransferase activity as compared with acyl-CoA synthetase activity was found in most tissue preparations, while the rate of the activating step matched that of the overall process. This suggests that formation of the acyl-CoA thioester (3PBA-CoA) is the rate-limiting step in the conjugation of 3PBA with glycine. 4. In most of the species tested, renal activities were higher than those of corresponding liver preparations. 5. The gerbil and ferret, which excrete 3-phenoxybenzoylglycine as the principal urinary metabolite of 3PBA, gave the highest 3PBA-CoA synthetase and glycine N-acyltransferase activities in vitro. By contrast, the hamster, which excretes only small amounts of the glycine conjugate of 3PBA, had the lowest enzymic activities in vitro. 6. In the mouse and rat there were differences between the patterns of metabolism found in vivo and in vitro, and possible reasons for this are discussed.

Acetate-CoA Ligase↗
PubMed1975

The identification and biosynthesis of siderochromes formed by Micrococcus denitrificans.

1. Micrococcus denitrificans excretes three catechol-containing compounds, which can bind iron, when grown aerobically and anaerobically in media deficient in iron, and anaerobically in medium with a high concentration of Ca2+. 2. One of these compounds was identified as 2,3-dihydroxybenzoic acid (compound I), and the other two were tentatively identified as N1N8-bis-(2,3-dihydroxybenzoyl)spermidine (compound II) and 2-hydroxybenzoyl-N-L-threonyl-N4[N1N8-bis-(2,3-dihydroxybenzoyl)]spermidine (compound III). 3. The equimolar ferric complex of compound III was prepared; compound III also forms complexes with Al3+, Cr3+ and Co2+ ions. 4. Cell-free extracts from iron-deficient organisms catalyse the formation of compound II from 2,3-dihydroxybenzoic acid and spermidine, and of compound III from compound II, L-threonine and 2-hydroxybenzoic acid; both reactions require ATP and dithiothreitol, and Mg2+ stimulates activity. The enzyme system catalysing the formation of compound II has optimum activity at pH 8.8 Fe2+ (35muM), Fe3+ (35muM) and Al3+ (65muM) inhibit the reaction by 50 percent. The enzyme system forming compound III has optimum activity at pH 8.6. Fe2+ (110 muM), Fe3+ (110 muM) and Al3+ (135 muM) inhibit the reaction by 50 percent. 5. At least two proteins are required for the formation of compound II, and another two proteins for its conversion into compound III. 6. The changes in the activities of these two systems were followed after cultures became deficient in iron. 7. Ferrous 1,10-phenanthroline is formed when a cell-free extract from iron-deficient cells is incubated with the ferric complex of compound III, succinate, NADH and 1,10-phenanthroline under N2.

Adenosine Triphosphate↗
PubMed1973

Aminolaevulinate synthetase of Micrococcus denitrificans. Purification and properties of the enzyme, and the effect of growth conditions on the enzyme activity in cells.

1. 5-Aminolaevulinate synthetase was detected in extracts of the non-photosynthetic bacterium Micrococcus denitrificans. 2. Activity is high in cells grown anaerobically in a defined nitrate medium, but is low in cells grown in an iron-deficient medium, and in cells grown aerobically. 3. Aminolaevulinate synthetase was purified extensively; it has a molecular weight of about 68000; apparent K(m) values for glycine, succinyl-CoA and pyridoxal phosphate are 12mm, 10mum and 11mum respectively; 2mum-haemin and 14mum-protoporphyrin inhibit by 50%. 4. The low activity of aminolaevulinate synthetase in iron-deficient cells increases on adding iron salts to cells only under conditions where protein synthesis can occur. 5. In defined nitrate medium with a high Ca(2+) concentration anaerobic growth yield is higher, but aminolaevulinate synthetase activity is lower than in cells grown in the medium with a low Ca(2+) concentration. In medium made from AnalaR constituents, growth yield is low and aminolaevulinate synthetase activity is high even in the presence of high concentrations of Ca(2+); on adding Cu(2+) (0.1mum) to the medium growth yield and aminolaevulinate synthetase activity become the same as in non-AnalaR medium. 6. Cells incubated under conditions where protein synthesis does not occur but where electron transport does, lose their aminolaevulinate synthetase activity rapidly. 7. The activities of aminolaevulinate dehydratase and succinic thiokinase do not change under any of the conditions of growth examined. 8. The possible mechanisms controlling aminolaevulinate synthetase activity and the role of this enzyme in controlling the synthesis of haem in this organism are discussed.

5-Aminolevulinate Synthetase↗
PubMed1973

Control of 5-aminolaevulinate synthetase activity in Rhodopseudomonas spheroides. The involvement of sulphur metabolism.

1. The ;initial' 5-aminolaevulinate synthetase activity, that is the activity observed immediately after cell disruption, in extracts prepared from unharvested semianaerobically grown Rhodopseudomonas spheroides, was twice that observed under the same assay conditions in extracts prepared from harvested cells. 2. The effect of oxygenation of a culture on the ;maximum' aminolaevulinate synthetase activity, that is the activity observed 1h after disruption of harvested cells, is markedly influenced by the contents of the growth medium. Oxygenation of organisms for 1h in the medium in which they have grown produces an 80-90% decrease in maximum activity, whereas similar treatment of organisms resuspended in fresh medium produces less than a 40% decrease. 3. This protective effect of fresh medium is absolutely dependent on the presence of sulphate. When cells are suspended in sulphate-deficient fresh medium, the maximum activity falls by 65-75% even without oxygenation. A high maximum activity is regenerated when sulphate is resupplied. 4. When organisms are oxygenated in the medium in which they have grown, the cellular contents of GSH+GSSG and cysteine+cystine fall very markedly and homolanthionine is formed. Both the fall in aminolaevulinate synthetase activity and the changes in sulphur metabolism are largely prevented by the addition of compounds which stimulate synthesis of cysteine de novo or inhibit the conversion of cysteine S into homocysteine S. 5. The maximum aminolaevulinate synthetase activity was directly proportional to the GSH+GSSG content of all cell preparations. In glutathione-depleted extracts the ;low'-activity enzyme could be re-activated in vitro by the addition of GSH, GSSG, cysteine or cystine, whereas in extracts with a high glutathione content the ;high'-activity enzyme was unaffected by these sulphur compounds. 6. The activation of low-activity enzyme with exogenous sulphur compounds was prevented by excluding air or by adding NADH. Studies with purified enzyme indicate that sulphur compounds do not interact directly with the enzyme, but that their effect is mediated by a number of other endogenous factors.

5-Aminolevulinate Synthetase↗
PubMed1973

Control of 5-aminolaevulinate synthetase activity in Rhodopseudomonas spheroides. The purification and properties of an endogenous activator of the enzyme.

1. A low-molecular-weight activator of 5-aminolaevulinate synthetase was detected in extracts of Rhodopseudomonas spheroides. The compound activates the enzyme extracted from oxygenated semi-anaerobically grown organisms by a factor of 6-8. 2. The activator was extensively purified, but owing to the exceedingly small amounts that could be extracted in the active form its structure was not determined. 3. The activator contains an acetylatable amino group; it is more stable at acid than at alkaline pH values; it is stable to treatment with I(2)-KI or potassium ferricyanide, but irreversibly inactivated by Na(2)S(2)O(4) or NaBH(4). 4. The chromatographic, electrophoretic, chemical and stability properties of the activator are similar to those of pteridines; purified activator preparations contain pteridines, as shown by their fluorescence spectrum. This does not, however, constitute an identification of the activator. 5. The activator enhances the activity of crude and partially purified enzyme and does not appear to require other endogenous factors or a supply of air to produce activation. Activation of the purified enzyme, however, requires the presence of either pyridoxal phosphate or sodium succinate. In the absence of both these factors the activator produces a time- and temperature-dependent decay of activity.

5-Aminolevulinate Synthetase↗
PubMed1972

Coproporphyrinogenase activities in extracts of Rhodopseudomonas spheroides and Chromatium strain D.

1. The anaerobic coproporphyrinogenase activity in an extract of Rhodopseudomonas spheroides is inhibited by 1,10-phenanthroline, alphaalpha'-bipyridyl, flavins, 2,4-dinitrophenol and 1,4-naphthaquinone. These compounds have no effect on the aerobic coproporphyrinogenase activity. 2. On removal of small-molecular-weight material from a crude extract, the anaerobic system becomes very unstable; it can be stabilized by adding succinate. Now nicotinamide nucleotides, in addition to Mg(2+), ATP and methionine, are required for protoporphyrin to be formed. 3. A mechanism for the anaerobic reaction is proposed, based on the cofactor requirements and the effect of inhibitors. 4. The enzyme responsible for aerobic activity has been partially purified and some of its properties are reported. 5. A crude extract of Chromatium strain D also exhibits coproporphyrinogenase activity under anaerobic conditions in the presence of S-adenosylmethionine or ATP plus methionine. The requirement for other cofactors is variable.

Adenosine Triphosphate↗
PubMed1970

Activation of delta-aminolaevulate synthetase in extracts of Rhodopseudomonas spheroides.

1. The spontaneous activation of delta-aminolaevulate synthetase in extracts from Rhodopseudomonas spheroides grown semi-anaerobically in the light requires oxygen and does not take place anaerobically in the dark. 2. Activation is completely prevented by azide or cyanide and is partially inhibited by chlorpromazine. These compounds inhibit markedly the succinoxidase activity of extracts. 3. NADH delays activation, but when it has been oxidized by the extract activation begins at the normal rate and complete activation occurs. By contrast both the rate and the extent of activation are markedly decreased by even small amounts of carboxylic acids. 4. The inhibitory effects of succinate and citrate on activation can be prevented by malonate and fluorocitrate respectively. 5. These results suggest that for activation to occur some endogenous compound has to be oxidized via the electron transport chain. 6. Activation occurs under anaerobic conditions in the light, probably by photo-oxidation.

Amino Acids↗