Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

---

Query: EC:2.4.99.6 (sialyltransferase)
1,546 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The beta-galactoside alpha-2,6-sialyltransferase is a trans Golgi/trans Golgi network glycosyltransferase which adds sialic acid residues to Asn-linked oligosaccharides of glycoproteins. Previous results suggested that the sialyltransferase stem and signal anchor including flanking sequences may be two independent Golgi retention regions. However, other experiments demonstrated that the sequence of the signal anchor itself was not important. To investigate whether the sialyltransferase signal anchor was necessary and sufficient for Golgi retention, several mutant and chimeric proteins were expressed and localized in Cos-1 and Chinese hamster ovary cells. We found that the signal anchor and flanking sequences were able to retain the sialyltransferase catalytic domain in the Golgi. However, efficient Golgi retention was still observed when the signal anchor was altered or entirely replaced in either the presence or absence of most of the luminal stem region. Chimeric proteins consisting of the sialyltransferase cytoplasmic tail and signal anchor fused to the extracellular domains of two different cell surface proteins demonstrated poor Golgi retention. A significant increase in the Golgi retention of one of these chimeras was observed when two lysines were placed next to the signal anchor on the luminal side. Taken together these results suggest that the sialyltransferase signal anchor is not necessary or sufficient for Golgi retention, rather, appropriately spaced cytoplasmic and luminal flanking sequences are the important elements of the sialyltransferase Golgi retention region.
...
PMID:Specific sequences in the signal anchor of the beta-galactoside alpha-2,6-sialyltransferase are not essential for Golgi localization. Membrane flanking sequences may specify Golgi retention. 825 53

Spermatozoa acquire fertilizing ability during passage through the epididymis. Modification of oligosaccharide moieties on sperm surface glycoproteins are some of the biochemical changes believed to be important in the production of functionally mature spermatozoa during passage through the epididymis. In an attempt to understand the mechanism underlying these modifications, we quantified four glycosyltransferase activities (the enzymes that catalyze the transfer of sugar residues from nucleotide sugar donor to the sugar chains on glycoproteins and glycolipids) of spermatozoa and fluid from various regions of the epididymis. Our results are as follows. (1) Only 10-20% of the total glycosyltransferase activities (sialyltransferase, fucosyltransferase, galactosyltransferase, and N-acetyl glucosaminyltransferase) sedimented with the spermatozoa; the remaining 80-90% of the four enzymes were present in soluble form in the epididymal fluid. (2) When the four transferase activities were expressed per 10(6) spermatozoa, only sialyltransferase and fucosyltransferase activities showed maturation-dependent changes. The former enzyme was significantly higher on the proximal caput spermatozoa and the latter on the distal caput spermatozoa. The higher levels of the two enzymes on caput spermatozoa could be due to their binding to the endogenous sugar acceptor molecules on the sperm surface, and subsequent release following sequential sialylation and fucosylation of the molecules in the proximal and distal caput spermatozoa, respectively. (3) When spermatozoa from the proximal and distal caput, corpus, and proximal and distal cauda were incubated with fucose-labeled nucleotide sugar (GDP[14C]fucose), higher levels of radioactivity were routinely incorporated into the spermatozoa from the distal caput. (4) The [14C]fucose-labeled spermatozoa or sperm plasma membranes, when solubilized, resolved on SDS-PAGE, and visualized by autoradiography, showed that the radioactivity had been incorporated into an endogenous acceptor of 86 kDa (major component) and several minor components. Treatment of the solubilized spermatozoa with N-glycanase suggested that the [14C]fucose is mainly present on N-linked oligosaccharide units. These studies demonstrate that some of the sperm surface components are fucosylated during sperm maturation. The potential significance of the in vitro fucosylation of sperm surface components in the production of functionally mature spermatozoa is discussed.
...
PMID:Glycosylation of rat sperm plasma membrane during epididymal maturation. 843 31

Mammalian spermatozoa must undergo maturational changes between the events of mating and fertilization. These biochemical and functional alterations, collectively termed capacitation, take place as spermatozoa traverse the female reproductive tract. The preparatory biochemical changes include removal, modification, and reorganization of sperm surface molecules. Although details of all the changes are not known, lectin binding studies have provided evidence suggesting that carbohydrate moieties of sperm surface glycoproteins are modified during capacitation. In an attempt to gain insight into the potential modifications of sperm plasma membrane glycoproteins, we quantified glycoprotein-modifying enzyme activities in the uterine and oviductal fluid of the hamster during the 4 days of the estrous cycle. These enzymes are known to modify existing glycoproteins, either by adding sugar residues (glycosyltransferases) or by removing terminal sugar residues (glycosidases). Data from these studies showed that 1) levels of all glycosyltransferase activities assayed (sialyltransferase, fucosyltransferase, galactosyltransferase, and N-acetylglucosaminyltransferase) were negligible in the uterine fluid at the onset of ovulation (Day 1) but sharply increased preceding ovulation (Day 4); 2) levels of the four glycosyltransferase activities assayed were higher in the oviductal fluid at the onset of ovulation (Day 1) and then gradually decreased through the remainder of the estrous cycle (Day 2 to Day 4); 3) levels of all glycohydrolase activities (acidic alpha-D-mannosidase, beta-D-galactosidase, beta-D-glucuronidase, beta-D-glucosaminidase, and alpha-L-fucosidase) and protein in the uterine and oviductal fluids did not vary widely during the 4 days of the cycle. These results demonstrate a temporal surge of glycosyltransferase activities in the genital tract fluids of the hamster. The temporal changes in the glycoprotein-modifying enzymes may have an effect on the glycosylation of sperm plasma membrane and zona pellucida glycoproteins at the site of fertilization or may alter the surface glycoproteins of the fertilized egg in the uterus prior to implantation.
...
PMID:Temporal surge of glycosyltransferase activities in the genital tract of the hamster during the estrous cycle. 872 23

Our goal was to engineer a Golgi glycosyltransferase epitope-tagged on its cytoplasmically exposed, short, N-terminal domain that gave normal subcellular localization. Partial replacement of the cytoplasmic tail of human alpha-2,6-sialyltransferase (SialylT) with the negatively charged myc or FLAG epitope resulted in almost complete mislocalization of the chimera expressed in Vero cells. A granular cytoplasmic staining pattern was seen by immunofluorescence. Spacing the negatively charged residues progressively outward from the negative N-terminus resulted in increasingly more normal localization of myc or FLAG-tagged protein to a juxtanuclear Golgi-like distribution. Substitution of a neutrally charged VSV-G sequence for these tags resulted in normal localization of the chimera to the juxtanuclear Golgi region. Insertion of the myc epitope within the N-terminal domain of the short form of bovine beta-1,4-galactosyltransferase (GalT) gave a chimeric protein that mislocalized in BHK cells. No signal was detected with a monoclonal anti-epitope antibody indicating that the myc epitope was masked. Placement of myc or FLAG epitopes at the NH2-terminus of human N-acetylglucosaminyltransferase I (GlcNAc-T) resulted in chimeric proteins that in Vero cells displayed little Golgi localization. We conclude that positioning of negative charge, in particular, close to the membrane, typically produces a failure of type II Golgi glycosyltransferases to exit the ER/CGN, presumably due to quality control mechanisms. These proteins may be successfully epitope-tagged on their N-terminal domain either using a neutral or positively charged sequence or spacing any negatively charged sequence out from the membrane.
...
PMID:Modification of the cytoplasmic domain affects the subcellular localization of Golgi glycosyl-transferases. 888 78

The genes encoding the alpha-2,3-sialyltransferases involved in lipooligosaccharide biosynthesis from Neisseria meningitidis and Neisseria gonorrhoeae have been cloned and expressed in Escherichia coli. A high sensitivity enzyme assay using a synthetic fluorescent glycosyltransferase acceptor and capillary electrophoresis was used to screen a genomic library of N. meningitidis MC58 L3 in a "divide and conquer" strategy. The gene, denoted lst, was found on a 2. 0-kilobase fragment of DNA, and its sequence was determined and then used to design probes to amplify and subsequently clone the corresponding lst genes from N. meningitidis 406Y L3, N. meningitidis M982B L7, and N. gonorrhoeae F62. Functional sialyltransferase was produced from the genes derived from both L3 N. meningitidis strains and the N. gonorrhoeae F62. However, the N. meningitidis M982B L7 gene contained a frameshift mutation that renders it inactive. The expression of the lst gene was easily detected using the enzyme assay, and the protein expression could be detected when an immunodetection tag was added to the COOH-terminal end of the protein. Using the synthetic acceptor N-acetyllactosamine-aminophenyl-(6-(5-(fluorescein-carboxamido)-hexan oic acid amide), the alpha-2,3 specificity of the enzyme was confirmed by NMR examination of the reaction product. The enzyme could also use synthetic acceptors with lactose or galactose as the saccharide portion. This study is the first example of the cloning, expression, and examination of alpha-2,3-sialyltransferase activity from a bacterial source.
...
PMID:Cloning of the lipooligosaccharide alpha-2,3-sialyltransferase from the bacterial pathogens Neisseria meningitidis and Neisseria gonorrhoeae. 891 Apr 46

Intracellular glycosyltransferase protein expression can be assessed by flow cytometry. We report the detectability of the Golgi associated beta1,4 galactosyltransferase (GT) and alpha2,6 sialyltransferase (ST) upon permeabilization in Jurkat and EBV-JY cells representing a T- and B-lymphoid cell line, respectively. The method employs fixation with paraformaldehyde and permeabilization with saponin. It ensures reliable internalization of the antibody and little background staining and does not cause leakage of the antigen. We first applied monoclonal antibodies to GT for establishment of this method by flow cytometry. The obtained flow cytometric signal could be localized to the Golgi apparatus by confocal laser scanning microscopy. To exclude interference from possible cell-surface staining, measurements were carried out on non-permeabilized cells. No signal was found in Jurkat cells, while low but measurable ecto-galactosyltransferase was found on EBV-JY cells. F(ab)'2 fragments of polyclonal antisera to GT and ST were generated and shown to be useful for double indirect staining with the monoclonal antibody. The method described here permits relative assessment of Golgi glycosyltransferase expression in lymphoid cells by flow cytometry.
...
PMID:Flow cytometric detection of the Golgi apparatus using antibodies to glycosyltransferases. 913 55

Human colorectal cancers express various cancer-associated carbohydrate determinants such as Lewis Y or sialyl Lewis A, suggesting a considerable alteration in glycosyltransferase activities occurring upon malignant transformation. We investigated the mRNA amounts of fucosyltransferase (Fuc-T) and sialyltransferase (ST) isoenzymes, including Fuc-T III, IV, V, VI and VII and ST-3N, ST-30 and ST-4, in human colorectal cancer tissues by Northern blotting and RT-PCR. Regarding fucosyltransferases, mRNA of Fuc-T III and VI was not significantly altered, and only Fuc-T IV mRNA showed a moderate increase in cancer tissues when compared with adjacent non-malignant colonic epithelia taken from the same patient (273 +/- 96%; p < 0.001). The moderate increase of Fuc-T IV message may be related to an enhanced expression of Lewis Y in colon cancer tissues. In the ST isoenzymes, mRNA for ST-3N remained unchanged, whereas that for ST-4 decreased significantly in cancer tissues, to 32 +/- 29%, (p < 0.005). The most remarkable finding was that the message of ST-30 was prominently increased in cancer tissues compared with non-malignant colorectal mucosa. When further investigated by quantitative RT-PCR assays on a larger series of patients with colorectal cancers, the average increase in mRNA for ST-30 was 459 +/- 200% compared with that in adjacent non-malignant epithelium (significant at p < 0.0001). The increase of ST-30 message was more prominent in the cancer tissues strongly expressing sialyl Lewis A than in the cancer tissues expressing sialyl Lewis A only weakly or moderately (significant at p < 0.05). The marked increase in the message of ST-30 is suggested to be related to an enhanced expression of sialylated carbohydrate determinants in colon cancer tissues including sialyl Lewis A, since the enzyme exhibited a significant activity against the type 1 chain carbohydrate substrate and produced the precursors for sialyl Lewis A synthesis, when its cDNA was expressed in Cos-7 cells.
...
PMID:Altered mRNA expression of specific molecular species of fucosyl- and sialyl-transferases in human colorectal cancer tissues. 917 8

The alpha2,3 sialyltransferase, alpha2,3 SAT (O), catalyzes the transfer of sialic acid to Galbeta1,3 N-acetyl-D-galactosamine (GalNAc) (core-1) in mucin type O-glycosylation, and thus terminates chain extension. A Core-2 branch can also be formed from core-1 by the core-2 beta1,6 N-acetyl-d-glucosamine transferase (beta1,6 GlcNAc T) that leads to chain extension. Increased levels of the alpha2,3 SAT (O) and decreased levels of the core-2 beta1,6 GlcNAc T are seen in breast cancer cells and correlate with differences in the structure of the O-glycans synthesized (Brockhausen et al., 1995; Lloyd et al., 1996). Since in mucin type O-glycosylation sugars are added individually and sequentially in the Golgi apparatus, the position of the transferases, as well as their activity, can determine the final structure of the O-glycans synthesized. A cDNA coding for the human alpha2,3 SAT (O) tagged with an immunoreactive epitope from the myc gene has been used to map the position of the glycosyltransferase in nontumorigenic (MTSV1-7) and malignant (T47D) breast epithelial cell lines. Transfectants were analyzed for expression of the enzyme at the level of message and protein, as well as for enzymic activity. In T47D cells, which do not express core-2 beta1,6 GlcNAc T, the increased activity of the sialyltransferase correlated with increased sialylation of core-1 O-glycans on the epithelial mucin MUC1. Furthermore, in MTSV1-7 cells, which do express core-2 beta1,6 GlcNAc T, an increase in sialylated core-1 structures is accompanied by a reduction in the ratio of GlcNAc: GalNAc in the O-glycans attached to MUC1, implying a decrease in branching. Using quantitative immunoelectron microscopy, the sialyltransferase was mapped to the medial- and trans-Golgi cisternae, with some being present in the TGN. The data represent the first fine mapping of a sialyltransferase specifically active in O-glycosylation and demonstrate that the structure of O-glycans synthesized by a cell can be manipulated by transfecting with recombinant glycosyltransferases.
...
PMID:A transfected sialyltransferase that is elevated in breast cancer and localizes to the medial/trans-Golgi apparatus inhibits the development of core-2-based O-glycans. 918 58

Mucin type O-glycans with core 2 branches are distinct from nonbranched O-glycans, and the amount of core 2 branched O-glycans changes dramatically during T cell differentiation. This oligosaccharide is synthesized only when core 2 beta-1, 6-N-acetylglucosaminyltransferase (C2GnT) is present, and the expression of this glycosyltransferase is highly regulated. To understand how O-glycan synthesis is regulated by the orderly appearance of glycosyltransferases that form core 2 branched O-glycans, the subcellular localization of C2GnT was determined by using antibodies generated that are specific to C2GnT. The studies using confocal light microscopy demonstrated that C2GnT was localized mainly in cis to medial-cisternae of the Golgi. We then converted C2GnT to a trans-Golgi enzyme by replacing its Golgi retention signal with that of alpha-2,6-sialyltransferase, which resides in trans-Golgi. Chinese hamster ovary cells expressing wild type C2GnT and the chimeric C2GnT were then subjected to oligosaccharide analysis. The results obtained clearly indicate that the conversion of C2GnT into a trans-Golgi enzyme resulted in a substantial decrease of core 2 branched oligosaccharides. These results, taken together, strongly suggest that the predominance of core 2 branched oligosaccharides in those cells expressing C2GnT is due to the fact that C2GnT is located earlier in the Golgi than alpha-2,3-sialyltransferase that competes with C2GnT for the common substrate. Furthermore, alteration of Golgi localization renders the chimeric C2GnT much less efficient in synthesizing core 2 branched oligosaccharides, indicating the critical role of orderly subcellular localization of glycosyltransferases.
...
PMID:Altered Golgi localization of core 2 beta-1,6-N-acetylglucosaminyltransferase leads to decreased synthesis of branched O-glycans. 927 27

Sialyl-Lex (sLex) antigen expression recognized by KM93 monoclonal antibody was significantly down-regulated during differentiation induced by 12-O-tetradecanoylphorbol-13-acetate (TPA) in human pre-B lymphocytic leukemia cell line KM3. The sLex determinants were almost exclusively expressed on O-linked oligosaccharide chains of an O-glycosylated 150-kDa glycoprotein (gp150). A low shear force cell adhesion assay showed that TPA treatment significantly inhibited E-selectin-mediated cell adhesion. Transcript and/or enzyme activity levels of alpha1-->3-fucosyltransferase, alpha2-->3-sialyltransferase, beta1-->4-galactosyltransferase, and elongation beta1-->3-N-acetylglucosaminyltransferase did not correlate with sLex expression levels. However, transcript and enzyme activity levels of core 2 GlcNAc-transferase (C2GnT) were significantly down-regulated during TPA treatment. Following transfection and constitutive expression of full-length exogenous C2GnT transcript, C2GnT enzyme activities were maintained at high levels even after TPA treatment and down-regulation of cell surface sLex antigen expression by TPA was completely abolished. Furthermore, in the transfected cells, the KM93 reactivity of gp150 was not reduced by TPA treatment, and the inhibition of cell adhesion by TPA was also blocked. These results suggest that sLex expression is critically regulated by a single glycosyltransferase, C2GnT, during differentiation of KM3 cells.
...
PMID:Single glycosyltransferase, core 2 beta1-->6-N-acetylglucosaminyltransferase, regulates cell surface sialyl-Lex expression level in human pre-B lymphocytic leukemia cell line KM3 treated with phorbolester. 975 22


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>

---