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:3.2.1.26 (invertase)
4,927 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have mapped a sequence determinant in the vacuolar glycoprotein carboxypeptidase Y (CPY) that directs intracellular sorting of this enzyme. Through the study of hybrid proteins, consisting of amino-terminal segments of CPY fused to the secretory enzyme invertase, we have found that the N-terminal 50 amino acids of CPY are sufficient to direct delivery of a CPY-Inv hybrid protein to the yeast vacuole. Our data suggest that this 50 amino acid segment of CPY contains two distinct functional domains; an N-terminal signal peptide followed by a segment of 30 amino acids that contains the vacuolar sorting signal. Deletion of this putative vacuole sorting signal from an otherwise wild-type CPY protein leads to missorting of CPY. Furthermore, examination of the Asn-linked oligosaccharides present on CPY and CPY-Inv hybrid proteins suggests that an additional determinant in CPY specifies the extent to which these proteins are glycosylated in the Golgi complex.
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PMID:Distinct sequence determinants direct intracellular sorting and modification of a yeast vacuolar protease. 302 48

CaCo-2 cells are human colonic adenocarcinoma cells which can differentiate spontaneously into enterocytes when maintained confluent for extended periods of time. Cells kept in culture for 4 days (rapidly growing), 7-9 days (early confluence) and 19-22 days (late confluence) were incubated for 24 h with L-[5,6-3H]fucose or D-[6-3H]glucosamine in order to examine the changes in glycoprotein carbohydrate structure that occur during this differentiation. Labelled glycopeptides obtained by exhaustive Pronase digestion of the cell-surface and cell-pellet fractions were fractionated on Bio-Gel P-6. A high-Mr glycopeptide fraction which was excluded from Bio-Gel P-6 was present in all cases. These glycopeptides were then fractionated by affinity chromatography on Datura stramonium agglutinin-agarose. The glycopeptides which were specifically bound to the lectin column were largely degraded by endo-beta-galactosidase, thereby indicating that they consisted of fucosylated polylactosaminoglycans. The proportion of labelled polylactosaminoglycans decreased with increasing time in culture, whereas sucrase activity, which is characteristic of differentiated enterocytes, increased. These results demonstrate that a relatively large decrease in the proportion of fucosylated polylactosaminoglycans occurs with differentiation of CaCo-2 cells.
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PMID:Differentiation-associated decrease in the proportion of fucosylated polylactosaminoglycans of CaCo-2 human colonic adenocarcinoma cells. 312 22

We studied lactase, maltase, and sucrase activities in the mucosa of self-filling blind loops (SFBL) in adult rats at weekly intervals after SFBL formation in order to determine the sequence in which disaccharidase activities fall. The studies were carried out on nourished and malnourished rats and extended to a recovery period induced by antibiotics to determine the effects of malnutrition on the establishment and repair of disaccharidase deficiencies caused by bacterial overgrowth. Malnutrition was produced by feeding 50% of the intake of paired rats fed ad libitum. Disaccharidase activities were determined in SFBL from nourished and malnourished rats at 7-day intervals until pandisaccharidase deficiency was established and during a 2-wk recovery period induced by antibiotics. Maximal SFBL bacterial counts in both nourished and malnourished groups of rats and brush-border glycoprotein degradation ratios were established at 7 days. In nourished rats only lactase was deficient at 7 days; maltase and sucrase fell later and sequentially. In malnourished rats all three disaccharidases were reduced at 7 days. Disaccharidase activities in self-emptying blind loops (SEBL), used as operated controls, were not decreased 28 days after surgery. Malnutrition had no effect on disaccharidase activities in the SEBL, and malnutrition did not affect recovery rates with antibiotic therapy. We conclude that small intestinal bacterial overgrowth causes a staggered loss of disaccharidase activities beginning with the loss of lactase activity. In the presence of bacterial overgrowth, malnutrition accelerates the conversion of a mono- to a pan-disaccharidase deficiency.
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PMID:Sequential disaccharidase loss in rat intestinal blind loops: impact of malnutrition. 315 66

Yeast invertase exists in two different forms. The cytoplasmic enzyme is nonglycosylated, whereas the external invertase contains about 50% carbohydrate of the high mannose type. The protein moieties of both enzymes are identical. The two invertases have been used previously as a model system to investigate the influence of covalently linked carbohydrate chains on the stability of large glycoproteins, and controversial results were obtained. Here, we measured thermal and denaturant-induced unfolding by various probes, such as the loss of enzymatic activity, and by the changes in absorbance and fluorescence. The ranges of stability of the two invertases were found to be essentially identical, indicating that the presence of a high amount of carbohydrate does not significantly contribute to the stability of external invertase. Earlier findings that invertase is stabilized by glycosylation could not be confirmed. The stability of this glycoprotein is apparently determined by the specific interactions of the folded polypeptide chain. Unlike the glycosylated form, the carbohydrate-free invertase is prone to aggregation in the denatured state at high temperature and in a partially unfolded form in the presence of intermediate concentrations of guanidinium chloride.
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PMID:The stability of yeast invertase is not significantly influenced by glycosylation. 328 23

Yeast secretory mutant sec53 cells accumulate inactive secretory glycoprotein precursors that remain associated with the endoplasmic reticulum (ER) at the restrictive temperature (37 degrees C). The possibility that precursor polypeptides fail to penetrate completely into the ER lumen was tested by examining the protease accessibility of accumulated invertase, mating pheromone precursor prepro-alpha-factor and the vacuolar protein precursor procarboxypeptidase Y in cell lysates. In all three cases, the secretory protein precursors are protected from the action of exogenous protease unless the membrane is permeabilized by including Triton X-100 or saponin in the incubation. These results suggest that the sec53 defect allows complete polypeptide translocation. Consistent with this interpretation, the precursor of invertase accumulates in a signal peptide-processed form. In addition, invertase and prepro-alpha-factor precursors contain a small amount of possibly aberrant carbohydrate. In mutant cells or in wild type cells treated with tunicamycin, a 10-kDa fragment of the N terminus of mature invertase assumes a conformation that is resistant to trypsin with or without detergent. This domain may be associated with an ER protein or may simply assume an unusual conformation as a consequence of deficient glycosyl modification.
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PMID:Product of SEC53 is required for folding and glycosylation of secretory proteins in the lumen of the yeast endoplasmic reticulum. 329 55

We have developed a purification procedure for the isolation of constitutive post-Golgi secretory vesicles from Saccharomyces cerevisiae. Although the post-Golgi stage of the secretion pathway is normally very rapid, we have used a temperature-sensitive secretory mutant, sec 6-4, to greatly expand the population of secretory vesicles. Following invertase as a marker, intact vesicles are enriched 36-fold from the crude lysate. The final preparation contains few contaminants as assessed by morphologic and biochemical examination. Three proteins (110, 40-45, and 18 kD) co-purify with the vesicle marker enzyme invertase. Metabolic labeling experiments indicate that these vesicle-associated proteins are synthesized during the period of vesicle accumulation. They are not apparent in the corresponding fractions from wild-type cells. Analysis of these proteins indicates that the 110-kD protein is a major glycoprotein residing in the vesicle lumen, while the 40-45- and 18-kD proteins are not glycosylated and are firmly associated with the vesicle membrane, each with at least one domain exposed on the cytoplasmic surface.
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PMID:Purification and characterization of constitutive secretory vesicles from yeast. 330 65

A mutant of Saccharomyces cerevisiae with the genotype mnn1 mnn2 mnn9 gls1 synthesizes mannoproteins with oligosaccharides having the composition Glc3Man10Glc-NAc2- owing to the mnn9 defect, which prevents synthesis of the outer chain, the mnn1 defect, which prevents branching of the core, and the gls1 mutation, which prevents deglucosylation of the resultant glycoprotein as a consequence of a defective glucosidase-I [Tsai, P.-K., Ballou, L., Esmon, B., Schekman, R. & Ballou, C. E. (1984) Proc. Natl. Acad. Sci. USA 81, 6340-6343]. (The mnn2 defect is not expressed in presence of the mnn9 mutation.) This strain spontaneously forms new colonies in which gls1 is suppressed owing to a defect in synthesis of dolichol phosphoglucose, the glucosylation substrate. The new mutant, designated mnn1 mnn2 mnn9 gls1 dpg1, synthesizes and secretes invertase (EC 3.2.1.26) that has a higher mobility on native gel electrophoresis than that made by the parent strain, the consequence of a reduction in both the size and the number of carbohydrate chains. The mannoprotein chains have the mnn1 mnn9 structure (Man10Glc-NAc2-), and the invertase is resolved by gel electrophoresis in sodium dodecyl sulfate into two major and two minor bands that represent homologs with about 4-7 carbohydrate units, in contrast to about 8-11 chains in the parent strain. Thus, the inability to glucosylate the lipid-linked precursor reduces the efficiency of glycosylation of the protein chains. The genetic defect is in synthesis of the glucose donor dolichol phosphoglucose, but the mutation is nonallelic with the reported alg5-1 mutation, which has a similar phenotype [Runge, K. W., Huffaker, T. C. & Robbins, P. W. (1984) J. Biol. Chem. 259, 412-417].
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PMID:A mutation that prevents glucosylation of the lipid-linked oligosaccharide precursor leads to underglycosylation of secreted yeast invertase. 351 49

A series of high mannose oligosaccharides with the size range Man8-14GlcNAc was purified from Saccharomyces cerevisiae invertase, and the composition of each was determined by chemical analysis. Purity and composition were verified by 1H NMR spectroscopy at 500 MHz, and structures were assigned on the basis of chemical shifts in C1-H and C2-H protons of similarly substituted compounds of known structure. Such analyses showed that these invertase oligosaccharides were a homologous series of homogeneous compounds, each related to the next member by addition of 1 mol of mannose in a specific alpha-linked configuration. Man8GlcNAc purified from the total glycoprotein fraction of disrupted yeast was the smallest species found and had the same homogeneous structure as that previously reported for the Man8GlcNAc from invertase (Byrd, J. C., Tarentino, A. L., Maley, F., Atkinson, P. H., and Trimble, R. B. (1982) J. Biol. Chem. 257, 14657-14666). Digestion of Man8-13GlcNAc species from invertase with Aspergillus satoi alpha 1,2-mannosidase provided products that were consistent with the structures assigned by 1H NMR as did fast atom bombardment-mass spectroscopy fragmentation analysis of the Man9,10GlcNAc oligosaccharides. These results lead to the proposal that Man8GlcNAc is the only trimming intermediate in Saccharomyces sp., and the remaining Man9-14GlcNAc oligosaccharides are biosynthetic intermediates which define the principal pathway of single-step mannose addition in the formation of the inner core of yeast mannan.
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PMID:Structure of yeast external invertase Man8-14GlcNAc processing intermediates by 500-megahertz 1H NMR spectroscopy. 352 34

We have devised a genetic selection for mutant yeast cells that fail to properly deliver the vacuolar glycoprotein CPY to the lysosome-like vacuole. This has allowed us to identify mutations in eight VPL complementation groups that result in aberrant secretion of up to approximately 90% of the immunoreactive CPY. Other soluble vacuolar proteins are also affected by each vpl mutation, demonstrating that a sorting system for multiple vacuolar proteins exists in yeast. Mislocalized CPY apparently traverses late stages of the secretory pathway, since a vesicle-accumulating sec1 mutation prevents secretion of this protein. Despite the presence of abnormal membrane-enclosed organelles in some of the vpl mutants, maturation and secretion of invertase are not substantially perturbed. Thus vpl mutations define a new class of genes that encode products required for sorting of newly synthesized vacuolar proteins from secretory proteins during their transit through the yeast secretory pathway.
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PMID:Protein sorting in yeast: mutants defective in vacuole biogenesis mislocalize vacuolar proteins into the late secretory pathway. 353 26

Tunicamycin apparently inhibited the biosynthesis of glucose, galactose, and maltose transport systems in Saccharomyces cerevisiae. Under the conditions used, the antibiotic also blocked the biosynthesis of invertase, a well-known yeast glycoprotein, as well as the glycosylation of a marker mannoprotein of the yeast cell wall. However, the antibiotic did not affect certain proteins which did not contain carbohydrate. It seems, therefore, that these sugar carriers are glycoproteins.
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PMID:Inhibition of biosynthesis of Saccharomyces cerevisiae sugar transport system by tunicamycin. 353 86


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