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

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

This review summarizes data on the distribution of certain glycosylation steps in the Golgi apparatus as revealed by immunolabeling and lectin techniques. The methodical basis for such investigations was provided by the introduction of the colloidal gold marker system for immunolabeling and the development of new means of tissue processing such as the low-temperature embedding technique using Lowicryl K4M. The application of these techniques together with highly specific antibodies has provided much of the basis for our current understanding of the Golgi apparatus in functional terms. Thus, in many cell types, three Golgi apparatus compartments can be distinguished, whereas in others no such functional subdivision is evident. Investigations on sialyltransferase distribution have also provided direct evidence that GERL is structurally and functionally part of the Golgi apparatus.
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PMID:Localization of glycosylation sites in the Golgi apparatus using immunolabeling and cytochemistry. 182 23

In a previous work we found that human colorectal cancer tissues express increased levels of an alpha 2,6 sialyltransferase (alpha 2,6 ST) acting on N-acetyllactosaminic sequences (E.C. 2.4.99.1). In this study we have taken advantage of the known specificity of elderberry bark lectin (Sambucus nigra agglutinin, SNA) for NeuAc alpha 2, 6Gal/GalNAc structures to investigate the relationship between expression of alpha 2,6 sialyltransferase activity and occurrence of alpha 2,6-sialylated oligosaccharide sequences in human colorectal cancer cell lines. Three cell lines with opposite adhesion properties were used in this study: SW 948 cells grow adherent to the culture flask surface and express very low levels of enzyme activity; COLO 205 cells grow in non-adherent form and express the highest levels of alpha 2,6 ST activity; A non-adherent subline of SW 948 cells (SW 948 FL) was isolated and found to express high levels of alpha 2,6 ST activity. By using SNA-Sepharose chromatography we found that expression of alpha 2,6 ST activity correlates with the extent of alpha 2,6-sialylation of N-linked chains of glycoproteins but not with the presence of alpha 2,6-sialylated glycolipids.
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PMID:Alpha 2,6 sialylation of N-acetyllactosaminic sequences in human colorectal cancer cell lines. Relationship with non-adherent growth. 189 84

The Golgi complex is composed of at least four distinct compartments, termed the cis-, medial, and trans-Golgi cisternae and the trans-Golgi network (TGN). It has recently been reported that the organization of the Golgi complex is disrupted in cells treated with the fungal metabolite, brefeldin-A. Under these conditions, it was shown that resident enzymes of the cis-, medial, and trans-Golgi return to the ER. We report here that 300-kD mannose 6-phosphate receptors, when pulse-labeled within the ER of brefeldin-A-treated cells, acquired numerous N-linked galactose residues with a half time of approximately 2 h, as measured by their ability to bind to RCA-I lectin affinity columns. In contrast, Limax flavus lectin chromatography revealed that less than 10% of these receptors acquired sialic acid after 8 h in brefeldin-A. Two lines of evidence suggested that proteins within and beyond the TGN did not return to the ER in the presence of brefeldin-A. First, the majority of 300-kD mannose 6-phosphate receptors present in the TGN and endosomes did not return to the ER after up to 6 h in brefeldin-A, as determined by their failure to contact galactosyltransferase that had relocated there. Moreover, although mannose 6-phosphate receptors did not acquire sialic acid when present in the ER of brefeldin-A-treated cells, they were readily sialylated when labeled at the cell surface and transported to the TGN. These experiments indicate that galactosyltransferase, a trans-Golgi enzyme, returns to the endoplasmic reticulum in the presence of brefeldin-A, while the bulk of sialyltransferase, a resident of the TGN, does not. Our findings support the proposal that the TGN is a distinct, fourth compartment of the Golgi apparatus that is insensitive to brefeldin-A.
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PMID:Compartmentation of the Golgi complex: brefeldin-A distinguishes trans-Golgi cisternae from the trans-Golgi network. 216 98

1. Subcellular fractions isolated from livers of 19-day-old chicken embryos were analyzed in order to assess whether liver mitochondria contained glycosylated proteins or had mannosyl- or sialyl-transferases that could transfer sugars to mitochondrial macromolecules. 2. Proteins in liver mitochondrial membranes and matrix fractions were screened for their affinities for concanavalin A (Con A). 3. After separation by gel electrophoresis under denaturing conditions, a significant number of the proteins bound [125I]Con A, and the binding of the lectin was substantially inhibited by alpha-methyl-D-mannoside. 4. In addition, radio-iodinated matrix proteins were screened for lectin-binding properties by chromatography on Con A covalently linked to agarose. 5. A number of proteins, representing 14% of those loaded onto the column, became tightly bound to the agarose-linked lectin, and the molecular weights of several of those proteins are reported. 6. Mannosyltransferase activities were measured in fractions highly enriched for mitochondria. 7. In the reactions, mannose was transferred from guanosine diphosphomannose to materials insoluble in 0.3% trichloroacetic acid or in chloroform:methanol (2:1). 8. The fractions also catalyzed the transfer of mannose to materials extractable in chloroform:methanol and which migrated with the Rf of dolichol phosphate on Silica Gel H. 9. Dolichol phosphate stimulated the transfer of mannose to those materials extractable in the organic solvents. 10. Marker enzyme analyses indicated that the mannosyl transferase activity in the mitochondrial fraction could not be accounted for entirely by contaminating microsomal membranes. 11. Although sialyltransferase activity was detected also in the mitochondrial fractions, the levels of the activity and the kinetics of the reactions indicated that Golgi membranes were most likely the sources of the enzyme.
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PMID:Mitochondrial biogenesis: do liver mitochondria contain glycoproteins and glycosyltransferases? 228 16

Alterations in cell surface proteins and glycoproteins may play a key role in determining the metastatic behavior of tumor cells. The cell surface proteins of a series of related murine colon cancer cells selected in an animal model for colon cancer metastasis (R. S. Bresalier et al., Cancer Res., 47: 1398-1406, 1987) were therefore compared by a variety of biochemical methods. Lactoperoxidase-catalyzed iodination of cell surface proteins followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis demonstrated quantitative and qualitative differences in the cell surface protein profiles of parental cell line 51B (low metastatic potential) and its metastatic derivatives 51B LiM 5 and 51B LiM 6. Labeling of sialic acid-containing proteins suggested that, in the case of at least four of these proteins (Mr 170,000, 120,000, 95,000, and 55,000), this represented an increase in radioactive labeling of sialoglycoproteins from the metastatic lines. Affinity chromatography of solubilized 125I-labeled cell membrane proteins revealed a 2- to 3-fold increase in wheat germ agglutinin and Sambucus nigra lectin binding associated with the metastatic lines, compared to the poorly metastatic parent. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of material eluted from these columns demonstrated enhancement of proteins from the metastatic cells corresponding in molecular weight to the previously identified major sialoglycoproteins. Neuraminidase-releasable membrane-associated sialic acid and sialyltransferase activities were 2- to 3-fold higher in the metastatic cell lines compared to the parental line. Liver colonization after intrasplenic injection of the various lines into syngeneic mice was dramatically reduced by prior removal of cell surface sialic acid. Immunohistochemical staining of primary and metastatic tumors formed after cecal injection of parental 51B suggested selective metastasis by wheat germ agglutinin-binding tumor cells. These results further support the concept that cell membrane sialylation is important in determining the metastatic potential of cancer cells.
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PMID:Cell surface sialoprotein alterations in metastatic murine colon cancer cell lines selected in an animal model for colon cancer metastasis. 229 75

Granulocytes from patients with chronic myelogenous leukemia (CML) are morphologically identical to their normal counterparts but show marked differences in circulation patterns and in some membrane properties. We have previously shown that there is abnormal lectin binding to CML granulocytes, and aberrant sialylation of membrane glycoproteins. To examine the changes in sialylation of CML granulocytes further, we have studied membrane preparations from CML and normal granulocytes for specific sialyltransferase activity. Because sialyltransferase enzymes are specific for the configuration of the acceptor group, enzyme activity was assayed by measuring transfer of sialic acid from CMP-14C-sialic acid to substrates of defined structure. As compared with those of normal counterparts, CML extracts catalyzed a 50% higher overall rate of sialylation of asialofetuin, a substrate possessing both N- and O-linked acceptors. Studies of enzyme specificity utilizing porcine and ovine submaxillary mucins, antifreeze glycoprotein and alpha-1 acid glycoprotein as acceptors showed that the increased sialylation by CML extracts was due primarily to substrates with the O-linked Gal beta 1----3GaINAc acceptor group. These data suggest that sialyltransferase activity is increased in CML granulocytes compared to normal granulocytes and that the increased enzyme activity is specific for O-linked Gal beta 1----3GaINAc. This enzyme activity may be directly responsible for the abnormal membrane sialylation and pathophysiological behavior of these cells.
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PMID:Increased activity of a specific sialyltransferase in chronic myelogenous leukemia. 241 27

The subcellular distribution of sialyltransferase and its product of action, sialic acid, was investigated in the undifferentiated cells of the rat intestinal crypts and compared with the pattern observed in the differentiated cells present in the surface epithelium. Sialyltransferase was immunocytochemically detected with an antibody, affinity-purified on a beta-galactosidase/sialyltransferase fusion protein, which recognizes only protein epitopes of the enzyme. A similar pattern and intensity of immunolabeling were observed in the Golgi apparatus, apical and basolateral plasma membranes of both undifferentiated and differentiated absorptive cells. However, in the goblet cells, the mucus was only weakly labeled in cells present in the basal portion of the crypts but increased in intensity through the zone of migration to the surface epithelium. Sialic acid as detected with the Limax flavus lectin was observed in the Golgi apparatus and post-Golgi apparatus structures of both absorptive and goblet cells regardless of their position along the crypt-to-surface epithelium axis. However, a striking difference in the plasma membrane distribution of sialic acid existed between undifferentiated cells of the lower half of the crypts and those of the upper half and the surface epithelium: in the former, label was present in both the apical and basolateral domain, whereas in the latter it became restricted to the apical domain. These results suggest that the presence of sialyltransferase immunoreactivity in the goblet cell mucus and the polarization of sialic acid to the apical plasma membrane of both goblet and absorptive cells may be markers for the differentiated state.
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PMID:Alteration in sialyltransferase and sialic acid expression accompanies cell differentiation in rat intestine. 245 28

Fluorescein isothiocyanate (FITC)-labelled asialotransferrin and pyridyl aminated oligosaccharides were prepared from asialotransferrin and human milk using affinity chromatography and high performance liquid chromatography (HPLC), respectively. These substances were incubated with galactosidase or sialyltransferase and then examined by lectin affinity HPLC. The elution patterns changed according to the period of incubation and amount of enzyme. This analytical method using lectin affinity HPLC with fluorescence labelled glycoprotein or oligosaccharides as the substrates has great value for detecting these enzyme under the same chromatographic conditions. In addition, differences were noted in the activity of beta-galactosidase toward oligosaccharides having the Gal beta(1----3)GlcNAc or Gal beta(1----4)GlcNAc structure at reducing termini.
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PMID:Rapid assay of beta-galactosidase and sialyltransferase by lectin affinity high performance liquid chromatography with fluorescence detection. 250 11

Sorting of newly synthesized proteins destined for the apical plasma membrane takes place in the trans-Golgi network (TGN) in MDCK cells. This process is most likely receptor mediated and requires components that recycle between both compartments. We have developed an assay to detect apical proteins that recycle through the sialyltransferase-containing TGN. Cell surface glycoproteins were exogalactosylated apically using a mutant cell line derived from MDCK, MDCKII-RCAr. The mutant exhibits impaired galactosylation of glycoconjugates and thereby allows maximal incorporation of exogenously added galactose in the presence of galactosyltransferase. Upon reculture at 37 degrees C, a time-dependent increase of sialylated apical surface glycoproteins was observed by lectin binding as well as by the sialic acid-specific NaIO4/NaB[3H]4 labeling technique. This indicates that some galactosylated surface molecules had returned to the TGN. Recycling through the TGN was blocked, if exogalactosylated cells were incubated at 20 degrees C. Two-dimensional gel electrophoresis identified three apical proteins which recycle through the TGN, suggesting that this pathway is selective for a subset of the apical surface proteins.
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PMID:A restricted set of apical proteins recycle through the trans-Golgi network in MDCK cells. 251 Sep 95

In this report we describe the alteration of the N-linked oligosaccharide terminal sequences of Chinese hamster ovary cell glycoproteins by expression of a beta-galactoside alpha 2,6-sialyltransferase cDNA. While wild type cells normally produce sugar chains terminating in the NeuAc alpha 2,3Gal linkage, the expressed enzyme competes with the endogenous sialyltransferase to attach an alternative terminal sequence, NeuAc alpha 2,6Gal. Subcellular localization of the NeuAc alpha 2,6Gal product by lectin-gold electron microscopy revealed localization throughout the Golgi apparatus cis to trans, post-Golgi membranes and vesicular structures. The results demonstrate the potential for purposefully altering terminal carbohydrate structures in vivo by "mis-expressing" terminal glycosyltransferases that compete with the endogenous enzyme normally produced by the cells.
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PMID:Alteration of terminal glycosylation sequences on N-linked oligosaccharides of Chinese hamster ovary cells by expression of beta-galactoside alpha 2,6-sialyltransferase. 266 74


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