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PubMed2003

Formyl-CoA transferase encloses the CoA binding site at the interface of an interlocked dimer.

Formyl-CoA transferase catalyses transfer of CoA from formate to oxalate in the first step of oxalate degradation by Oxalobacter formigenes, a bacterium present in the intestinal flora which is implicated in oxalate catabolism in mammals. Formyl-CoA transferase is a member of a family of CoA-transferases for which no structural information is available. We now report the three-dimensional structure of O.formigenes formyl-CoA transferase, which reveals a novel fold and a very striking assembly of the homodimer. The subunit is composed of a large and a small domain where residues from both the N- and C-termini of the subunit are part of the large domain. The linkers between the domains give the subunit a circular shape with a hole in the middle. The enzyme monomers are tightly interacting and are interlocked. This fold requires drastic rearrangement of approximately 75 residues at the C-terminus for formation of the dimer. The structure of a complex of formyl-CoA transferase with CoA is also reported and sets the scene for a mechanistic understanding of enzymes of this family of CoA-transferases.

Amino Acid Sequence↗
PubMed1993

Differences in induction by xenobiotics in murine tissues and the Hepa1c1c7 cell line of mRNAs encoding glutathione transferase, quinone reductase, and CYP1A P450s.

Levels of mRNAs encoding class-alpha glutathione transferases, class-mu glutathione transferases, quinone reductase, and cytochrome P450 1A were measured after xenobiotic induction in murine tissues and in the Hepa1c1c7 murine hepatoma cell line. RNA levels in liver and intestinal mucosa were determined after induction with phenobarbital, butylated hydroxyanisole, beta-naphthoflavone, isosafrole, or combinations of these compounds. The tissue culture cells were presented with combinations of butylated hydroxyanisole, tert-butyl-hydroquinone, and beta-naphthoflavone. In murine liver and intestinal mucosa, the greatest induction (5-15-fold) of glutathione transferases and quinone reductase was seen with butylated hydroxyanisole. Administration of phenobarbital or beta-naphthoflavone has only a modest effect (2-3-fold). In contrast, cytochrome P450 1A mRNA levels increase only slightly after BHA induction but are induced dramatically by beta-naphthoflavone. The pattern of induction is different in Hepa1c1c7 cells; there the greatest induction of all mRNAs occurred with beta-naphthoflavone. Administration of antioxidants with other xenobiotics increases mRNA levels only slightly over the levels obtained with BHA in murine tissues, or with beta-naphthoflavone in Hepa1c1c7 cells. mGSTM1 (GT8.7, Yb1), the most abundant glutathione transferase mRNA in murine liver, is also the most abundant glutathione transferase mRNA in both normal and induced Hepa1c1c7 cells. Our results suggest that BHA induction in murine liver and intestinal mucosa of class-mu and class-alpha glutathione transferases may involve regulatory elements and mediators that function poorly in Hepa1c1c7 cells.

Animals↗
PubMed1975

Multiple forms of human glutathione S-transferase and their affinity for bilirubin.

The initial enzymic step in mercapturic acid formation is catalyzed by glutathione S-transferase. Several species of this enzyme, designated as transferases alpha, beta, gamma, delta and epsilon on the basis of increasing isoelectric points, were isolated from human liver. Evidence is presented that each of the purified species is homogeneous with respect to sodium dodecylsulfate-gel electrophoresis. Transferases alpha, beta and epsilon each appear as a single band on gel electrofocusing; transferases gamma and delta are present as two and three bands, respectively, with each band catalytically active. Amino acid analysis indicated the five transferases to be either very closely related or identical in this respect. All enzyme species have a molecular weight of about 48500 and consist of two apparently identical subunits. The spectrum of substrates is the same for each although the enzymes differ slightly in specific activity. As is the case for the rat liver enzymes, each of the human transferases binds bilirubin although this compound is not a substrate.

Amino Acids↗
PubMed1981

Glutaconate CoA-transferase from Acidaminococcus fermentans.

1. Glutaconate CoA-transferase catalyses the transfer of CoAS- from acetyl-CoA preferentially to (E)-glutaconate, but glutarate, (R)-2-hydroxyglutarate, acrylate and propionate are also good acceptors. No reaction was observed with (Z)-glutaconate and C4-dicarboxylic acids. 2. The product of the reaction of acetyl-CoA with (E)-glutaconate is the 1-isomer of glutaconyl-CoA, i.e. the thiol ester is conjugated with the double bond. Other results indicate, however, that with (R)-2-hydroxyglutarate as substrate both possible isomers are generated. 3. Glutaconate CoA-transferase was purified from cell-free extracts of Acidaminococcus fermentans to apparent homogeneity and crystallized. The relative molecular mass of the enzyme is approximately 275000. It consists of two different polypeptide chains (M, 32000 and 34000). On the catalytic pathway a thiolester is formed between CoASH and a carboxylate of the smaller polypeptide chain. 4. The structural and functional relationships between glutaconate CoA-transferase and other CoA-transferases are discussed. 5. Glutaconate CoA-transferase is also present in other bacteria fermenting glutamate via hydroxyglutarate. Experiments with an antiserum against the enzyme indicate that the transferase is necessary for the decarboxylation of glutaconate but not for the dehydration of (R)-2-hydroxyglutarate.

Acyl Coenzyme A↗
PubMed1988

Purification and characterization of five forms of glutathione transferase from human uterus.

Five glutathione transferase (GST) forms were purified from human uterus by glutathione-affinity chromatography followed by chromatofocusing, and their structural, kinetic and immunological properties were investigated. Upon SDS/polyacrylamide slab gel electrophoresis all forms resulted composed of two subunits of identical molecular size. GST V (pI 4.5) is a dimer of 23-kDa subunits. GST I (pI 6.8) and GST IV (pI 4.9) are dimers of 24-kDa subunits whereas GST II (pI 6.1) and GST III (pI 5.5) are dimers of 26.5-kDa subunits. GST V accounts for about 85-90% of the activity whereas the other isoenzymes are present in trace quantities. On the basis of the molecular mass of the subunits, amino acid composition, substrate specificities, sensitivities to inhibitors, CD spectra and immunological studies, GST V appeared very similar to transferase pi. Structural and immunological studies provide evidence that GST IV is closely related to the less 'basic' transferase (GST pI 8.5) of human skin. Extensive similarities have been found between GST II and GST III. The comparison includes amino acid compositions, subunits molecular size and immunological properties. The two enzymes, however, are kinetically distinguishable. The data presented also indicate that GST II and GST III are related to transferase mu and to transferase psi of human liver. Even though GST I has a subunit molecular mass identical to GST IV, several lines of evidence, including catalytic and immunological properties, indicate that they are different from each other. GST I seems not to be related to any of known human transferases, suggesting that it may be specific for the uterus.

Affinity Labels↗
PubMed1994

Naturally occurring human glutathione S-transferase GSTP1-1 isoforms with isoleucine and valine in position 104 differ in enzymic properties.

Glutathione S-transferase P1-1 isoforms, differing in a single amino acid residue (Ile104 or Val104), have been previously identified in human placenta [Ahmad, H., Wilson, D. E., Fritz, R. R., Singh, S. V., Medh, R. D., Nagle, G. T., Awasthi, Y. C. & Kurosky, A. (1990) Arch. Biochem. Biophys. 278, 398-408]. In the present report, the enzymic properties of these two proteins are compared. [I104]glutathione S-transferase P1-1 has been expressed from its cDNA in Escherichia coli and purified to homogeneity by affinity chromatography; the cDNA has been mutated to replace Ile104 by Val104, and [V104]glutathione S-transferase P1-1 was expressed and isolated as described for [I104]glutathione S-transferase P1-1. The two enzymes differed in their specific activity and affinity for electrophilic substrates (KM values for 1-chloro-2,4-dinitrobenzene were 0.8 mM and 3.0 mM for [I-104]glutathione S-transferase P1-1 and [V-104]glutathione S-transferase P1-1, respectively), but were identical in their affinity for glutathione. In addition, the two enzymes were distinguishable by their heat stability, with half-lives at 45 degrees C of 19 min and 51 min, respectively. The resistance to heat denaturation was differentially modulated by the presence of substrates. These data, in conjunction with molecular modeling, indicate that the residue in position 104 helps to define the geometry of the hydrophobic substrate-binding site, and may also influence activity by interacting with residues directly involved in substrate binding.

Base Sequence↗
PubMed1996

Determination of a binding site for a non-substrate ligand in mammalian cytosolic glutathione S-transferases by means of fluorescence-resonance energy transfer.

To determine the location of the non-substrate-ligand-binding region in mammalian glutathione S-transferases, fluorescence-resonance energy transfer was used to calculate distances between tryptophan residues and protein-bound 8-anilinonaphthalene 1-sulphonate (an anionic ligand) in the human class-alpha glutathione S-transferase, and in a human Trp28-->Phe mutant class-pi glutathione S-transferase. Distance values of 2.21 nm and 1.82 nm were calculated for the class-alpha and class-pi enzymes, respectively. Since glutathione S-transferases bind one non-substrate ligand/protein dimer, the ligand-binding region, according to the calculated distances, is found to be located in the dimer interface near the twofold axis. This region is the same as that in which the parasitic helminth Schistosoma japonicum glutathione S-transferase binds praziquantel, a non-substrate drug used to treat schistosomiasis [McTigue, M. A., Williams, D. R. & Tainer, J. A. (1995) J. Mol. Biol. 246, 21-27]. Since the overall folding topology is conserved and certain features at the dimer interface are similar throughout the superfamily, it is reasonable to expect that all cytosolic glutathione S-transferases bind non-substrate ligands in the amphipathic groove at the dimer interface.

Binding Sites↗
PubMedDate not supplied

An extracellular monoADP-ribosyl transferase activity in Entamoeba histolytica trophozoites.

Due to the important role of monoADP-ribosyl transferases in physiological and pathological events, we investigated whether the protozoan parasite Entamoeba histolytica had monoADP-ribosyl transferase activity. Reactions were initiated using ameba-free medium as the source of both enzyme and ADP-ribosylation substrate(s) and [32P]NAD+ as source of ADP-ribose. Proteins were analyzed by electrophoresis, and [32P]-labeled proteins were detected by autoradiography. Using the crude extracellular medium, a major labeled product of Mr 37.000 was observed. The yield of this product was reduced markedly using medium from Brefeldin A-treated trophozoites, indicating that the extracellular monoADP-ribosyl transferase and/or its substrate depended on vesicular transport. The labeling of the 37-kDa substrate was dependent on reaction time, temperature, pH, and the ratio of unlabeled NAD+ to [32P]NAD+. After two purification steps, several new substrates were observed, perhaps due to their enrichment. The reaction measured ADP-ribosylation since [14C-carbonyl]NAD+ was not incorporated into ameba substrates and a 75-fold molar excess of ADP-ribose caused no detectable inhibition of the monoADP-ribosyl transferase reaction. On the basis of sensitivity to NH2OH, the extracellular monoADP-ribosyl transferase of E. histolytica may be an arginine-specific enzyme. These results demonstrate the existence in E. histolytica of at least one extracellular monoADP-ribosyl transferase, whose localization depends upon a secretion process.

ADP Ribose Transferases↗
PubMed1997

Effect of anaerobic environment on the glutathione transferase isoenzymatic pattern in Proteus mirabilis.

When Proteus mirabilis was cultured anaerobically in the presence of nitrate as terminal electron acceptor, a dramatic reduction of glutathione transferase production occurred. The analysis of the glutathione affinity purified materials in terms of substrate specificity, SDS-PAGE pattern, IEF pattern and immunoblotting revealed that a significantly different glutathione transferase pattern also occurred: two new glutathione transferase forms with an isoelectric point at pH 4.8 and 5.0 appeared. Their N-terminal amino acid sequence analysis as well as the ability to bind to a glutathione affinity column indicate that major differences between anaerobic and aerobic glutathione transferase forms are mainly located in the C-terminal region of the primary structure. In contrast, no significant changes occurred in the production of glutathione transferase isoenzymes when P. mirabilis was grown anaerobically in the absence of a terminal electron acceptor. These results support the idea that bacterial glutathione transferase expression is not strictly related to the absence of oxygen stress.

Anaerobiosis↗
PubMed1999

Variable expression of glutathione S-transferase isoenzymes in the fungus, Mucor circinelloides.

Mucor circinelloides (previously M. javanicus) variably expresses two isoenzymes of glutathione S-transferases. Glutathione S-transferases 1 and 2 were purified by affinity chromatography and found to be dimers with subunit M(r) values of 25.5 and 28. While glutathione S-transferase 1 immunoblotted with rat glutathione S-transferase T 5-5, N-terminal sequencing indicated that this enzyme was distinct from recently characterised bacterial and fungal glutathione S-transferases and did not readily fit into any one class. It showed some similarity to several classes including insect class II and plant type III. M. circinelloides glutathione S-transferase 2 did not immunoblot and yielded no N-terminal sequence.

Amino Acid Sequence↗
PubMed1990

Inverse relationship between total glutathione S-transferase content and bile acid release in isolated hepatocytes from untreated, phenobarbital pretreated and hypothyroid rats.

The role of cytosolic anion binding proteins (glutathione S-transferases) in the hepatic transport of bile acids remains controversial. To investigate whether increased levels of the hepatocyte total glutathione S-transferase content were associated with changes in the release of bile acids from the hepatocyte, we measured the rate of release of radioactive bile acids in isolated hepatocytes from thyroidectomized, phenobarbital pretreated and untreated rats. The isolated hepatocytes were preincubated with either 14C-cholic acid or 14C-taurocholic acid, and the release rate of radiolabeled bile acids was determined. Hepatocyte total glutathione S-transferase content was measured by rocket immunoelectrophoresis. The release rate of the radiolabeled bile acids was significantly (P less than 0.005) decreased in both hypothyroid and phenobarbital pretreated hepatocytes. The levels of total glutathione S-transferase content were significantly (P less than 0.001) increased in the hepatocytes from both hypothyroid and phenobarbital pretreated animals. Our findings reveal a striking inverse relationship between the total glutathione S-transferase content of the hepatocyte and the release rate of radiolabeled bile acids in isolated hepatocytes from two independent animal models. These observations support the hypothesis that cytosolic anion binding proteins (glutathione S-transferases) may influence the net flux across the hepatocyte plasma membrane largely by limiting efflux.

Animals↗
PubMed2006

Characterization of (R)-2-hydroxyisocaproate dehydrogenase and a family III coenzyme A transferase involved in reduction of L-leucine to isocaproate by Clostridium difficile.

The strictly anaerobic pathogenic bacterium Clostridium difficile occurs in the human gut and is able to thrive from fermentation of leucine. Thereby the amino acid is both oxidized to isovalerate plus CO(2) and reduced to isocaproate. In the reductive branch of this pathway, the dehydration of (R)-2-hydroxyisocaproyl-coenzyme A (CoA) to (E)-2-isocaprenoyl-CoA is probably catalyzed via radical intermediates. The dehydratase requires activation by an ATP-dependent one-electron transfer (J. Kim, D. Darley, and W. Buckel, FEBS J. 272:550-561, 2005). Prior to the dehydration, a dehydrogenase and a CoA transferase are supposed to be involved in the formation of (R)-2-hydroxyisocaproyl-CoA. Deduced amino acid sequences of ldhA and hadA from the genome of C. difficile showed high identities to d-lactate dehydrogenase and family III CoA transferase, respectively. Both putative genes encoding the dehydrogenase and CoA transferase were cloned and overexpressed in Escherichia coli; the recombinant Strep tag II fusion proteins were purified to homogeneity and characterized. The substrate specificity of the monomeric LdhA (36.5 kDa) indicated that 2-oxoisocaproate (K(m) = 68 muM, k(cat) = 31 s(-1)) and NADH were the native substrates. For the reverse reaction, the enzyme accepted (R)- but not (S)-2-hydroxyisocaproate and therefore was named (R)-2-hydroxyisocaproate dehydrogenase. HadA showed CoA transferase activity with (R)-2-hydroxyisocaproyl-CoA as a donor and isocaproate or (E)-2-isocaprenoate as an acceptor. By site-directed mutagenesis, the conserved D171 was identified as an essential catalytic residue probably involved in the formation of a mixed anhydride with the acyl group of the thioester substrate. However, neither hydroxylamine nor sodium borohydride, both of which are inactivators of the CoA transferase, modified this residue. The dehydrogenase and the CoA transferase fit well into the proposed pathway of leucine reduction to isocaproate.

Alcohol Oxidoreductases↗
PubMed2006

Properties of succinyl-coenzyme A:D-citramalate coenzyme A transferase and its role in the autotrophic 3-hydroxypropionate cycle of Chloroflexus aurantiacus.

The phototrophic bacterium Chloroflexus aurantiacus uses the 3-hydroxypropionate cycle for autotrophic CO(2) fixation. This cycle starts with acetyl-coenzyme A (CoA) and produces glyoxylate. Glyoxylate is an unconventional cell carbon precursor that needs special enzymes for assimilation. Glyoxylate is combined with propionyl-CoA to beta-methylmalyl-CoA, which is converted to citramalate. Cell extracts catalyzed the succinyl-CoA-dependent conversion of citramalate to acetyl-CoA and pyruvate, the central cell carbon precursor. This reaction is due to the combined action of enzymes that were upregulated during autotrophic growth, a coenzyme A transferase with the use of succinyl-CoA as the CoA donor and a lyase cleaving citramalyl-CoA to acetyl-CoA and pyruvate. Genomic analysis identified a gene coding for a putative coenzyme A transferase. The gene was heterologously expressed in Escherichia coli and shown to code for succinyl-CoA:d-citramalate coenzyme A transferase. This enzyme, which catalyzes the reaction d-citramalate + succinyl-CoA --> d-citramalyl-CoA + succinate, was purified and studied. It belongs to class III of the coenzyme A transferase enzyme family, with an aspartate residue in the active site. The homodimeric enzyme composed of 44-kDa subunits was specific for succinyl-CoA as a CoA donor but also accepted d-malate and itaconate instead of d-citramalate. The CoA transferase gene is part of a cluster of genes which are cotranscribed, including the gene for d-citramalyl-CoA lyase. It is proposed that the CoA transferase and the lyase catalyze the last two steps in the glyoxylate assimilation route.

Acyl Coenzyme A↗
PubMed2001

Succinyl-CoA:(R)-benzylsuccinate CoA-transferase: an enzyme of the anaerobic toluene catabolic pathway in denitrifying bacteria.

Anaerobic microbial toluene catabolism is initiated by addition of fumarate to the methyl group of toluene, yielding (R)-benzylsuccinate as first intermediate, which is further metabolized via beta-oxidation to benzoyl-coenzyme A (CoA) and succinyl-CoA. A specific succinyl-CoA:(R)-benzylsuccinate CoA-transferase activating (R)-benzylsuccinate to the CoA-thioester was purified and characterized from Thauera aromatica. The enzyme is fully reversible and forms exclusively the 2-(R)-benzylsuccinyl-CoA isomer. Only some close chemical analogs of the substrates are accepted by the enzyme: succinate was partially replaced by maleate or methylsuccinate, and (R)-benzylsuccinate was replaced by methylsuccinate, benzylmalonate, or phenylsuccinate. In contrast to all other known CoA-transferases, the enzyme consists of two subunits of similar amino acid sequences and similar sizes (44 and 45 kDa) in an alpha(2)beta(2) conformation. Identity of the subunits with the products of the previously identified toluene-induced bbsEF genes was confirmed by determination of the exact masses via electrospray-mass spectrometry. The deduced amino acid sequences resemble those of only two other characterized CoA-transferases, oxalyl-CoA:formate CoA-transferase and (E)-cinnamoyl-CoA:(R)-phenyllactate CoA-transferase, which represent a new family of CoA-transferases. As suggested by kinetic analysis, the reaction mechanism of enzymes of this family apparently involves formation of a ternary complex between the enzyme and the two substrates.

Acyl Coenzyme A↗
PubMed2002

Degradation of aromatics and chloroaromatics by Pseudomonas sp. strain B13: purification and characterization of 3-oxoadipate:succinyl-coenzyme A (CoA) transferase and 3-oxoadipyl-CoA thiolase.

The degradation of 3-oxoadipate in Pseudomonas sp. strain B13 was investigated and was shown to proceed through 3-oxoadipyl-coenzyme A (CoA) to give acetyl-CoA and succinyl-CoA. 3-Oxoadipate:succinyl-CoA transferase of strain B13 was purified by heat treatment and chromatography on phenyl-Sepharose, Mono-Q, and Superose 6 gels. Estimation of the native molecular mass gave a value of 115,000 +/- 5,000 Da with a Superose 12 column. Polyacrylamide gel electrophoresis under denaturing conditions resulted in two distinct bands of equal intensities. The subunit A and B values were 32,900 and 27,000 Da. Therefore it can be assumed that the enzyme is a heterotetramer of the type A2B2 with a molecular mass of 120,000 Da. The N-terminal amino acid sequences of both subunits are as follows: subunit A, AELLTLREAVERFVNDGTVALEGFTHLIPT; subunit B, SAYSTNEMMTVAAARRLKNGAVVFV. The pH optimum was 8.4. Km values were 0.4 and 0.2 mM for 3-oxoadipate and succinyl-CoA, respectively. Reversibility of the reaction with succinate was shown. The transferase of strain B13 failed to convert 2-chloro- and 2-methyl-3-oxoadipate. Some activity was observed with 4-methyl-3-oxoadipate. Even 2-oxoadipate and 3-oxoglutarate were shown to function as poor substrates of the transferase. 3-oxoadipyl-CoA thiolase was purified by chromatography on DEAE-Sepharose, blue 3GA, and reactive brown-agarose. Estimation of the native molecular mass gave 162,000 +/- 5,000 Da with a Superose 6 column. The molecular mass of the subunit of the denatured protein, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, was 42 kDa. On the basis of these results, 3-oxoadipyl-CoA thiolase should be a tetramer of the type A4. The N-terminal amino acid sequence of 3-oxoadipyl-CoA thiolase was determined to be SREVYI-DAVRTPIGRFG. The pH optimum was 7.8. Km values were 0.15 and 0.01 mM for 3-oxoadipyl-CoA and CoA, respectively. Sequence analysis of the thiolase terminus revealed high percentages of identity (70 to 85%) with thiolases of different functions. The N termini of the transferase subunits showed about 30 to 35% identical amino acids with the glutaconate-CoA transferase of an anaerobic bacterium but only an identity of 25% with the respective transferases of aromatic compound-degrading organisms was found.

Acetyl-CoA C-Acyltransferase↗
PubMedDate not supplied

Histidine 229 in protein L2 is apparently essential for 50S peptidyl transferase activity.

It has recently been suggested that peptidyl transferase activity is primarily a property of ribosomal RNA and that ribosomal proteins may act only as scaffolding. On the other hand, evidence from both photoaffinity labeling studies and reconstitution studies suggest that protein L2 may be functionally important for peptidyl transferase. In the work reported here, we reconstitute 50S subunits in which the H229Q variant of L2 replaces L2, with all other ribosomal components remaining unchanged, and determine the catalytic and structural properties of the reconstituted subunits. We observe that mutation of the highly conserved His 229 to Gin results in a complete loss of peptidyl transferase activity in the reconstituted 50S subunit. This is strong evidence for the direct involvement of L2 in ribosomal peptidyl transferase activity. Control experiments show that, though lacking peptidyl transferase activity, 50S subunits reconstituted with H229Q-L2 appear to be identical with 50S subunits reconstituted with wild-type L2 with respect to protein composition and 70S formation in the presence of added 30S subunits. Furthermore, as shown by chemical footprinting analysis, H229Q-L2 appears to bind 23S RNA in the same manner as wild-type L2. Thus, the effect of H229 mutation appears to be confined to an effect on peptidyl transferase activity, providing the most direct evidence for protein involvement in this function to date.

Amino Acid Sequence↗
PubMed1994

Quantitation of glutathione transferase-pi in the urine by radioimmunoassay.

A radioimmunoassay procedure for the quantitation of glutathione transferase-pi was developed in order to determine the levels of this protein in human urine. The enzyme was isolated from human placenta with a purification factor of 366 (compared to the original high-speed supernatant fraction), and upon gel electrophoresis, only a single band was seen. Polyclonal antisera were subsequently raised in rabbits and found to be suitable for a radioimmunoassay. Glutathione transferase-pi was localized immunohistochemically to the cells of the distal tubules, the thin loop of Henle and the collecting ducts in the kidney. In contrast, the alpha-isoenzyme was localized exclusively in the proximal tubular epithelium. Samples of urine from healthy individuals contained about 6 ng of the pi-transferase/ml. The method proved to be specific for glutathione transferase-pi, and no cross-reaction with the alpha- or mu-transferase or with other proteins occasionally appearing in urine occurred. The protein was quite stable upon storage and insensitive to variations in the urine pH. Thus, it appears that glutathione transferase-pi can be conveniently quantitated by radioimmunoassay and changes in the concentration of this protein in human urine thus monitored.

Enzyme Stability↗
PubMed1987

Developmental changes in intestinal glycosyl-transferase activities.

The carbohydrate composition of some intestinal glycoproteins has been demonstrated previously to be modified during development. To evaluate the role of the enzymatic mechanisms of glycosylation, the activities of three soluble and microsomal glycosyl-transferases were studied during postnatal development in rat intestinal mucosa. Nonexistent as soluble forms in the cell sap from suckling rats, the membrane-bound N-acetylgalactosaminyl-, sialyl-, and fucosyl-transferases showed different activities in microsomal fractions before weaning. The N-acetylgalactosaminyl-transferase activity was constant whereas the sialyl-transferase activity, which was maintained at a rather high level until the age of 2 wk, decreased just before weaning, while the fucosyl-transferase activity declined progressively from birth to weaning. After weaning, the three enzymatic activities progressively enhanced until adult age. Their variable behaviors on several glycoprotein acceptors may suggest the presence of several sialyl--and fucosyl-transferases with differences in specificities, varying in different ways according to age. Such developmental modifications in intestinal glycosylation patterns may be linked with certain transformation observed in the carbohydrate level of mucus or membrane-bound glycoproteins and related to the profound modifications in nutritional status at the weaning period.

Animals↗