Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.24.27 (thermolysin)
 1,894 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The dialdehyde produced by the periodate cleavage of the ribose moiety of uridine 5'-diphosphate (UDP) has been used as an affinity label for the UDP-galactose/UDP binding site of galactosyltransferase from bovine colostrum. This derivative causes progressive inactivation of galactosyltransferase at a rate dependent on its concentration, and under certain conditions is a competitive inhibitor with respect to UDP-galactose. The substrate UDP-galactose protects the enzyme from inactivation. The inactivation is also dependent on Mn2+ concentration in a range that implies that the binding of Mn2+ at site I is a prerequisite for the binding of the UDP derivative. The inactivation can be progressively reversed by nitrogenous bases, or stabilized by KBH4 reduction, which is consistent with the hypothesis that a Schiff base has formed with a lysine residue. Galactosyltransferase was inactivated with a [3H]UDP derivative and the predominant labeled peptide, from thermolysin digestion, isolated and characterized as: Ser-Gly-Lys-UDP.
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PMID:Affinity labeling of bovine colostrum galactosyltransferase with a uridine 5'-diphosphate derivative. 95 73

We investigated the asialo-agalactofetuin galactosyltransferase solubilized by Triton X-100 from a human bladder transitional cell carcinoma line (cell-associated form). The specific activity of this enzyme was dependent on cell population density, being about 50% higher in cells from confluent than from sparse cultures. We compared the properties of this enzyme with those of a galactosyltransferase isoenzyme present in the culture medium (soluble form). Electrophoresis on nondenaturing polyacrylamide gels showed the two forms to be isoenzymes, in that the mobility of the soluble enzyme was greater than that of the cell-associated enzyme. The isoenzymes differed in that the Km for UDP-galactose of the cell-associated enzyme (1 X 10(-5) M) was one-half that of the soluble isoenzyme. The isoenzymes differed by 1 order of magnitude in their affinity for a fetuin-derived acceptor with a Km of 16 X 10(-5) M for the cell-associated and 1.2 X 10(-5) M for the soluble form, although the Km for ovalbumin and asialomucin as acceptor was similar for both. Both enzymes were active over a broad pH range and their response to divalent cations was the same: the most efficient cation was Mn2+; but modest activity was detected in the presence of either Cd2+ or Co2+. As determined by gel filtration on Sepharose 6B, the cell-associated galactosyltransferase showed a molecular weight of 66,000, whereas that of the soluble form was 51,000. Limited proteolysis of the cell-associated enzyme with thermolysin and subsequent analysis by nondenaturing polyacrylamide gel electrophoresis demonstrated that the cell-associated enzyme could be converted to an isoenzyme showing the same electrophoretic mobility as the soluble enzyme present in the culture medium, presumably by removal of a portion of the peptide chain. The same result was obtained by treating the cell-associated enzyme with a cell extract. This suggests but does not prove that the soluble enzyme secreted or shed into the medium is produced from the cell-associated form by an endogenous protease.
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PMID:Comparison of cell-associated and soluble galactosyltransferase isoenzymes from a human bladder transitional cell carcinoma line. 310 74

As analyzed by a phase-separation technique, the Triton X-114 extract of human mammary carcinoma cells (MCF-7 cells) contain an amphiphilic form of galactosyltransferase (UDPgalactose: D-glucose 4-beta-D-galactosyltransferase, EC 2.4.1.22), while the galactosyltransferase activity released by these cells represents a hydrophilic form of the enzyme. When the amphiphilic galactosyltransferase was subjected to limited proteolysis with thermolysin, this treatment generated a hydrophilic form of the enzyme. With respect to Km for UDPgalactose the kinetic data were very similar for the amphiphilic, for the released and the hydrophilic galactosyltransferases produced by proteinase treatment. Differences were detected in electrophoretic and gel chromatographic properties. The hydrophilic enzymes showed a greater electrophoretic mobility on non-denaturing polyacrylamide gels than did the amphiphilic form. On Sepharose 6B column chromatography, the amphiphilic galactosyltransferase appeared to be of higher molecular weight than the hydrophilic enzyme.
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PMID:Conversion of the amphiphilic galactosyltransferase from human mammary carcinoma cells to an active hydrophilic enzyme form by limited proteolysis. 392 17

In the previous paper (Block, M. A., Dorne, A.-J., Joyard, J., and Douce, R. (1983) J. Biol. Chem. 258, 13273-13280), we have described a method for the separation of membrane fractions enriched in outer and inner envelope membranes from spinach chloroplasts. The two envelope membranes have a different weight ratio of acyl lipid to protein (2.5-3 for the outer envelope membrane and 0.8-1 for the inner envelope membrane). The two membranes also differ in their polar lipid composition. However, in order to prevent the functioning of the galactolipid:galactolipid galactosyltransferase during the course of envelope membrane separation, we have analyzed the polar lipid composition of each envelope membrane after thermolysin treatment of the intact chloroplasts. The outer envelope membrane is characterized by the presence of high amounts of phosphatidylcholine and digalactosyldiacylglycerol whereas the inner envelope membrane has a polar lipid composition almost identical with that of the thykaloids. No phosphatidylethanolamine or cardiolipin could be detected in either envelope membranes, thus demonstrating that the envelope membranes, and especially the outer membrane, do not resemble extrachloroplastic membranes. No striking differences were found in the fatty acid composition of the polar lipids from either the outer or the inner envelope membrane. The two envelope membranes also differ in their carotenoid composition. Among the different enzymatic activities associated with the chloroplast envelope, we have shown that the Mg2+-dependent ATPase, the UDP-Gal:diacylglycerol galactosyltransferase, the phosphatidic acid phosphatase, and the acyl-CoA thioesterase are associated with the inner envelope from spinach chloroplasts whereas the acyl-CoA synthetase is located on the outer envelope membrane.
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PMID:Preparation and characterization of membrane fractions enriched in outer and inner envelope membranes from spinach chloroplasts. II. Biochemical characterization. 663 Feb 30

Intact chloroplasts isolated from leaves of eight species of 16:3 and 18:3 plants and chromoplasts isolated from Narcissus pseudonarcissus L. flowers synthesize galactose-labeled mono-, di-, and trigalactosyldiacylglycerol (MGDG, DGDG, and TGDG) when incubated with UDP-[6-(3)H]galactose. In all plastids, galactolipid synthesis, and especially synthesis of DGDG and TGDG, is reduced by treatment of the organelles with the nonpenetrating protease thermolysin. Envelope membranes isolated from thermolysin-treated chloroplasts of Spinacia oleracea L. (16:3 plant) and Pisum sativum L. (18:3 plant) or membranes isolated from thermolysin-treated chromoplasts are strongly reduced in galactolipid:galactolipid galactosyltransferase activity, but not with regard to UDP-Gal:diacylglycerol galactosyltransferase. For the intact plastids, this indicates that thermolysin treatment specifically blocks DGDG (and TGDG) synthesis, whereas MGDG synthesis is not affected. Neither in chloroplast nor in chromoplast membranes is DGDG synthesis stimulated by UDP-Gal. DGDG synthesis in S. oleracea chloroplasts is not stimulated by nucleoside 5'-diphospho digalactosides. Therefore, galactolipid:galactolipid galactosyltransferase is so far the only detectable enzyme synthesizing DGDG. These results conclusively suggest that the latter enzyme is located in the outer envelope membrane of different types of plastids and has a general function in DGDG synthesis, both in 16:3 and 18:3 plants.
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PMID:Biosynthesis of digalactosyldiacylglycerol in plastids from 16:3 and 18:3 plants. 1666 15