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)

The inactivation of external yeast invertase by irradiation in dilute aqueous solution has been investigated. The contributions of the individual radical species from water radiolysis to inactivation and amino acid degradation were estimated from the results of experiments in which solutions were saturated with nitrogen, nitrous oxide or oxygen, and on addition of hydroxyl radical scavengers. Under conditions where inactivation by hydroxyl radicals predominates, the rate of inactivation increased with increasing dose, indicating that in the initial stages of the radiolysis the mannose-rich oligosaccharide chains of the glycoprotein protect the polypeptide chain from radical attack. Amino acid analysis of the irradiated external invertase showed that there was significant destruction of tyrosine, phenylalanine, methionine and histidine residues. Destruction of methionine and histidine residues may be responsible for the free radical-induced inactivation of this enzyme.
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PMID:The radiation-induced inactivation of external yeast invertase in dilute aqueous solution. 256 93

Synthetic oligonucleotides coding for the yeast invertase secretion signal peptide were fused to the gene for the mature form of human interferon (huIFN-alpha 2). Two plasmids (E3 and F2) were constructed. E3 contained the invertase signal codons in a reading frame with the mature huIFN-alpha 2 gene. F2 had a deletion of the codon for alanine at amino acid residue-5 in the invertase signal and an addition of a methionine codon located between the coding sequences for the invertase signal and mature huIFN-alpha 2. Both hybrid genes were located adjacent to the promoter from the 3-phosphoglycerate kinase gene on the multicopy yeast expression plasmid, YEp1PT. Yeast transformants containing these plasmids produced somewhat more IFN than did the same expression plasmid containing the IFN gene with its human secretion signal sequence. HuIFN-alpha 2, purified from the medium of yeast cells containing E3, was found to be processed at the correct site. The huIFN-alpha 2 made by plasmid F2 was found to be completely processed at the junction between the invertase signal (a variant) and the methionine of methionine-huIFN-alpha 2. These results strongly suggested that the invertase signal (or its variant) attached to huIFN was efficiently recognized by the presumed signal recognition particle and was cleaved by the signal peptidase in the yeast cells. These results also suggested that amino acid changes on the right side of the cleavage site did not necessarily prevent cleavage or secretion.
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PMID:Saccharomyces cerevisiae secretes and correctly processes human interferon hybrid proteins containing yeast invertase signal peptides. 302 6

Acid trehalase was purified from the yeast suc2 deletion mutant. After hydrophobic interaction chromatography, the enzyme could be purified to a single band or peak by a further step of either polyacrylamide gel electrophoresis, gel filtration, or isoelectric focusing. An apparent molecular mass of 218,000 Da was calculated from gel filtration. Polyacrylamide gel electrophoresis of the purified enzyme in the presence of sodium dodecyl sulfate suggested a molecular mass of 216,000 Da. Endoglycosidase H digestion of the purified enzyme resulted after sodium dodecyl sulfate gel electrophoresis in one distinct band at 41,000 Da, representing the mannose-free protein moiety of acid trehalase. The carbohydrate content of the enzyme was 86%. Amino acid analysis indicated 354 residues/molecule of enzyme including 9 cysteine moieties and only 1 methionine. The isoelectric point of the enzyme was estimated by gel electrofocusing to be approximately 4.7. The catalytic activity showed a maximum at pH 4.5. The activity of the enzyme was not inhibited by 10 mM each of HgCl2, EDTA, iodoacetic acid, phenanthrolinium chloride or phenylmethylsulfonyl fluoride. There was no activation by divalent metal ions. The acid trehalase exhibited an apparent Km for trehalose of 4.7 +/- 0.1 mM and a Vmax of 99 mumol of trehalose min-1 X mg-1 at 37 degrees C and pH 4.5. The acid trehalase is located in the vacuoles. The rabbit antiserum raised against acid trehalase exhibited strong cross-reaction with purified invertase. These cross-reactions were removed by affinity chromatography using invertase coupled to CNBr-activated Sepharose 4B. Precipitation of acid trehalase activity was observed with the purified antiserum.
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PMID:Purification and characterization of acid trehalase from the yeast suc2 mutant. 328 51

The effects of an oral neomycin and penicillin regimen on intestinal bacteriology and on morphology and function of the small intestine of mice were investigated. Quantitative and qualitative stool cultures on selective media of the treated animals revealed only growth of yeast organisms. The treated animals developed enlargement of the ceca with fluid contents and watery stools, resembling characteristics of germfree animals. Radioautography with tritiated thymidine revealed an increased epithelial cell migration rate in the mice treated with the antibiotics for 3 to 5 wk. A slight increase in villus height was also noted. The treated male mice showed greater variance than the treated females in epithelial cell migration rates. Histochemical staining reactions showed a decrease in nonspecific esterase and in NADH dehydrogenase activity in the proximal gut of the antibiotic animals. Stains of distal gut and those for acid and alkaline phosphatase, NADPH dehydrogenase, lactic dehydrogenase, and succinic dehydrogenase were similar to the controls. A slight increase in sucrase activity and a slight decrease in lactase activity in the antibiotic animals was observed in contrast to control animals. Germfree mice, however, had greater sucrase and lactase activity. Transport of L-methionine was slightly reduced in the distal segment of the treated animals. Since the direction of these changes is away from the intestinal state observed in germfree animals, they are probably the result of the direct action of the antibiotics on the gut.
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PMID:Effects of neomycin and penicillin administration on mucosal proliferation of the mouse small intestine. With morphological and functional correlations. 438 18

Rats were made severely uremic with partial nephrectomy (24-hour creatinine clearance 10% of normal). Jejunal dipeptidase activities (substrates: glycyl-L-leucine, L-alanyl-L-proline, and L-methionyl-L-methionine), disaccharidase activities (maltase, sucrase, trehalase, and lactase) and morphology were studied. A highly significant increase in glycyl-L-leucine and L-methionyl-L-methionine dipeptidases was found in uremic rats compared with controls. Proline dipeptidase activities were unaltered. Disaccharidase activities showed a slight increase in sucrase in uremic rats; otherwise no change was found.
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PMID:Small intestinal dipeptidases and disaccharidases in experimental uremia in rats. 677 73

Saccharomyces cerevisiae strain FH4C carries a single invertase structural gene, SUC2, but produces distinct invertase mRNAs and polypeptides for the secreted and cytoplasmic forms of the enzyme. The two major invertase cell-free translation products are polypeptides of 60,000 daltons (p60) and 62,000 daltons (p62) and correspond to the nonglycosylated cytoplasmic form of invertase and the precursor of glycosylated secreted invertase, respectively. This paper describes amino acid sequence and peptide map analyses of invertase polypeptides. The peptide maps demonstrate that p62, p60, and the in vivo secreted polypeptide have significant structural homology. Sequence analysis, however, revealed differences between p62 and p60 at their amino termini. p62 contains an amino-terminal signal sequence of 19 amino acid residues that is specifically cleaved during secretion in a cell-free system to generate the secreted 87,000-dalton invertase glycopeptide gp87. This signal sequence is not present in p60. p60 synthesis begins with a methionine which can be aligned with a methionine at residue 21 in p 62. During translation, the p60 initiator methionine is removed and the newly generated amino terminus is acetylated. Based on peptide map similarities, partial amino-terminal sequence data, and common genetic origin, it is suggested that p60 and p62 have identical amino acid sequences carboxy-terminal to the p60 initiator methionine (residue 21 of p62). The reciprocal correlations of signal sequence with secretion and absence of signal sequence with cytoplasmic localization provide proof of the signal hypothesis for secreted proteins. Two mechanisms are proposed for the derivation of p60 and p62 from a single structural gene: alternative promoter sites, and differential processing of a single primary transcript.
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PMID:Presecretory and cytoplasmic invertase polypeptides encoded by distinct mRNAs derived from the same structural gene differ by a signal sequence. 703 84

Explants of pig small intestine were maintained at 37 degrees C in organ culture for periods up to 24 h in a system using Trowell T-8 medium supplemented with 10% foetal-calf serum. The mucosal morphology was well preserved during culture, as judged by light and electron microscopy. The explant contents of protein and two brush-border enzymes, microvillus aminopeptidase (EC 3.4.11.2) and dipeptidyl peptidase IV (EC 3.4.14.5), were not significantly modified during culture compared with controls, but a moderate, continuous release of both protein and enzyme activities into the medium was observed. Continuous labelling with [35S]methionine resulted in an even incorporation of radioactivity in the protein components, and the rate of labelling only moderately decreased over the 24 h period. The polypeptide compositions of sucrase (EC 3.2.1.48)--isomaltase (EC 3.2.1.10), maltase--glucoamylase (EC 3.2.1.20) lactase (EC 3.2.1.23)--phlorizin hydrolase (EC 3.2.1.62), microvillus aminopeptidase and aspartate aminopeptidase (EC 3.4.11.7) synthesized during culture were studied, and some were found to be similar to those of the pro-forms of the enzymes isolated from animals that had had their pancreatic duct disconnected 3 days before being killed. These results confirmed earlier findings of the existence of pro-forms of some of the microvillar enzymes and thus indicate a low activity of pancreatic proteinases in the culture system.
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PMID:Biosynthesis of intestinal microvillar proteins. Characterization of intestinal explants in organ culture and evidence for the existence of pro-forms of the microvillar enzymes. 709 36

The development and distribution of peptidase activity in mucosal homogenates of rat small intestine has been investigated. Substrates used were glycyl-L-leucine (GL), L-seryl-L-methionine (SM), and L-leucyl-glycyl-glycine (LGG). During the first 2 weeks of life there was high peptidase activity toward GL and SM in the distal regions of the small intestine. In the third postnatal week, activity in the distal small intestine toward GL and SM decreased, while activity in the proximal small intestine increased. In contrast, there was no difference in activity toward LGG along the length of the small intestine, nor was there a developmental change. Activity toward all three substrates was not affected by cortisone acetate treatment. However, the classical effect of glucocorticoids on sucrase activity and body weight was observed. All peptidases studied showed maximal activities at neutral pH, indicating that they were not lysosomal in origin. Activity towards GL and SM was predominantly located in the cytosol. It is suggested that these dipeptidase activities play a role in the terminal steps of protein digestion following pinocytosis and lysosomal hydrolysis.
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PMID:Postnatal development of peptidase enzymes in rat small intestine. 718 19

To investigate the role of post-translational events in intestinal cell differentiation we have studied the effects of heat shock on processing and cell surface delivery of sucrase-isomaltase (SI), dipeptidylpeptidase IV (DPPIV) and aminopeptidase N (APN) in Caco-2 cells. In cells cultured at 42.5 degrees C there was a rapid decline in sucrase activity, while DPPIV and APN were unaffected over a 3-day period. Immunofluorescence staining confirmed the selective disappearance of SI from the surface membrane after only 1 day of culture at 42.5 degrees C. Cell-surface biotinylation of cells metabolically labelled with [35S]methionine 4 h after a switch from 37 degrees C to 42.5 degrees C demonstrated that newly synthesized APN and DPPIV were associated with the surface membrane, while SI was almost completely retained intracellularly. Pulse-chase experiments confirmed that, in these cells, DPPIV and APN were normally processed and vectorially delivered to the cell surface; in contrast, conversion between the two conformationally distinct high-mannose precursor forms of SI (hmP1 and hmP2) was markedly inhibited, a significant fraction of newly synthesized enzyme was degraded, probably in the ER, and an immature form of complex-glycosylated SI precursor (cP) was produced and mostly retained intracellularly. Double labelling of Caco-2 cells for SI and cathepsin D excluded an accumulation of SI in the lysosomes, suggesting that this organelle was not involved in the degradation of SI. These results indicate that the ER may play an important role in intestinal cell differentiation by regulating the conformational maturation, degradation and eventual cellular localization of some digestive enzymes.
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PMID:Intracellular degradation and reduced cell-surface expression of sucrase-isomaltase in heat-shocked Caco-2 cells. 810 Apr 14

We have previously isolated glycopeptides derived from yeast invertase that acted as highly potent elicitors in suspension-cultured tomato cells, inducing ethylene biosynthesis and phenylalanine ammonia-lyase activity, and we have found that the high mannose oligosaccharides released from the pure glycopeptide elicitors by endo-beta-N-acetylglucosaminidase H acted as suppressors of elicitor activity (Basse, C. W., Bock, K., and Boller, T. (1992) J. Biol. Chem. 267, 10258-10265). One of the elicitor-active glycopeptides (gp 8c) was labeled with t-butoxycarbonyl-L-[35S]methionine and purified by reversed phase high performance liquid chromatography resulting in a specific radioactivity of the derivative of about 900 Ci/mmol. This radiolabeled glycopeptide showed specific, saturable, and reversible binding to whole tomato cells under conditions in which cells are responsive to elicitors as well as to microsomal membranes derived from these cells. Ligand saturation experiments, performed with microsomal membranes, gave a dissociation constant (Kd) of 3.3 nM as determined by Scatchard analysis. Various glycopeptide elicitors and preparations from yeast invertase were compared with respect to their abilities to compete for binding of 35S-labeled gp 8c to tomato membranes and to induce ethylene biosynthesis in tomato cells. These studies revealed a high degree of correlation between elicitor activities in vivo and displacement activities in vitro. In both tests, a high activity depended on the presence of glycan side chains consisting of more than 8 mannosyl residues. The high mannose oligosaccharides that acted as suppressors of elicitor activity in vivo competed for binding of the labeled elicitor also. The suppressor-active glycan Man11GlcNAc and the elicitor-active gp 8c exhibited very similar displacement activities, and the inhibitory constant (Ki) of the glycan Man11GlcNAc was very similar to the Kd value calculated for 35S-labeled gp 8c, indicating that the glycopeptide elicitors and the glycan suppressors derived from these elicitors competed with similar affinities for the same binding site. The suppressor-inactive glycan Man8GlcNAc had a 200-fold lower capacity to compete for binding of 35S-labeled gp 8c to tomato membranes compared with the suppressor-active glycan Man11GlcNAc. Our results demonstrate the existence of a specific elicitor binding site in tomato cell membranes and suggest that glycopeptides and glycans act as agonists and antagonists for induction of the stress response, respectively, by competing for this binding site.
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PMID:High affinity binding of a glycopeptide elicitor to tomato cells and microsomal membranes and displacement by specific glycan suppressors. 832 50


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