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:3.4.21.4 (trypsin)
42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The digestive juice of Achatina balteata, a giant snail of the West African Coast catalyses the hydrolysis of several natural and synthetic compounds. Enzymatic activities on lactose, o- and p-nitrophenyl-beta-D-galactoside, p-nitrophenyl-beta-D-glucoside, p-nitrophenyl-beta-D (and alpha-L-) fucoside, o-nitrophenyl-beta-D-xyloside, p-nitrophenyl-N-acetyl-beta-D-glucosaminide and phenolphthalein-glucuronide have been shown to be present. The effect of pH and substrate concentration on these activities were studied. The galactosidase, glucosidase and fucosidase activities were studied with respect to temperature, heat inactivation, pH stability and incubation with trypsin. Kinetic experiments suggest the presence of several galactosidase activities. This hypothesis is confirmed by specific staining after polyacrylamide gel electrophoresis. These activities showed a broad specificity towards galactosides and glucosides. The digestive juice showed no action on acetyl-L-tyrosine and benzoyl-L-arginine ethyl esters. However a small protease activity was observed on hemoglobine. No lipase activity was found. Sulfatase content was low compared to that of Helix pomatia.
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PMID:[Characterization of some hydrolase activities in digestive juice of Achatina balteata]. 0 61

The predominant surface glycopeptide from a clone of baby hamster kidney cells transformed by Rous sarcoma virus (C13/B4), metabolically labeled with L-[14C]fucose, has been characterized for the first time. This glycopeptide represents 19% of the total radioactivity removed by trypsin from the cell surface of the transformed fibroblasts and is more abundant in the transformed cells than in the normal counterpart. Purification of the glycopeptide after digestion with Pronase was by successive chromatography on DEAE-cellulose and Sephadex G-50. The monosaccharide content of the glycopeptide was 42, 127, 138, 114, and 243 nmol of fucose, sialic acid, galatose, mannose, and glucosamine, respectively. A partial structure of the glycopeptide was proposed from the results of sequential enzymatic degradation coupled with gas-liquid chromatographic analysis of the resultant monosaccharides. All of the enzymes used were purified and pretested on natural substrates and found to remove terminal monosaccharides of the correct configuration, quantitatively. The purification and properties of an alpha-L-fucosidase from rat testes were described. All of the radioactivity in the glycopeptide, recovered as fucose, was present at the core and was removed by treatment with this alpha-L-fucosidase. The proposed structure is a triantennary, completely sialylated, complex glycopeptide containing a core region of beta-D-mannose, beta-D-N-acetylglucosamine, and alpha-L-fucose.
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PMID:Partial structure of a membrane glycopeptide from virus-transformed hamster cells. 22 Oct 11

Murine resident macrophages express, on their surface, carbohydrate epitopes which undergo changes during their stimulation/activation as monitored by binding of 125I labelled Evonymus europaea and Griffonia simplicifolia I-B4 lectins. Treatment of the stimulated macrophages with coffee bean alpha-galactosidase abolished binding of the GS I-B4 isolectin and changed the binding pattern of the Evonymus lectin. The affinity (Ka) of Evonymus lectin for alpha-galactosidase-treated macrophages decreased approximately 23-fold, from 1.25 x 10(8) M-1 to 5.5 x 10(6) M-1. Subsequent digestion of alpha-galactosidase-treated macrophages with alpha-L-fucosidase from Trichomonas foetus, further reduced binding of Evonymus lectin. Resident macrophages showed the same pattern of Evonymus lectin binding, with the same affinity, as alpha-galactosidase-treated, stimulated macrophages. These results, together with a consideration of the carbohydrate binding specificity of the Evonymus lectin which, in the absence of alpha-D-galactosyl groups, requires alpha-L-fucosyl groups for binding, indicate the presence, on resident macrophages, of glycoconjugates with terminal alpha-L-fucosyl residues. It is also concluded that during macrophage stimulation/activation alpha-D-galactosyl residues are added to this glycoconjugate and that they form part of the receptor for Evonymus lectin. The same glycoconjugate(s) is/are also expressed on the activated macrophage IC-21 cell line which exhibits the same characteristics as that of stimulated peritoneal macrophages, i.e., it contains alpha-D-galactosyl end groups and is resistant to the action of trypsin. Both lectins were also specifically bound to Corynaebacterium parvum activated macrophages.
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PMID:Alpha-D-galactosylation of surface fucoglycoconjugate(s) upon stimulation/activation of murine peritoneal macrophages. 134 14

Chromatographic methods were developed for the separation and characterization of acidic (sialylated) and neutral (asialo-complex and high-mannose) oligosaccharides released from glycoproteins with peptide N-glycosidase F. endo-beta-N-acetylglucosaminidase F and endo-beta-N-acetylglucosaminidase H using a carbohydrate analyzer (Dionex BioLC). All the carbohydrate separations were carried out on a polymeric pellicular anion-exchange column HPIC-AS6/CarboPac PA-1 (Dionex) using only two eluants namely, 0.5 M NaOH and 3% acetic acid/NaOH pH 5.5, which were mixed with water to generate various gradients. Developed conditions for quantitative detection of carbohydrates with pulsed amperometry were necessary to obtain steady baselines at 0.1-0.3 microA output with suitable sensitivity (less than 5 pmol) in separations employing a variety of acidic and alkaline sodium acetate gradients. Oligosaccharides released from heat-denatured and trypsin-treated glycoproteins were purified initially from large-scale digestion (greater than 0.1 g) by extraction of peptide material into phenol/chloroform and finally by ion-exchange chromatography of the acqueous phase. Oligosaccharides isolated from the peptide N-glycosidase digests of bovine fetuin, human transferrin and alpha 1-acid glycoprotein gave multiple peaks in each charge group in separations based on the charge content at pH 5.5. Alkaline sodium acetate gradients were developed to obtain oligosaccharide maps of the glycoproteins within 60 min, in which separated oligosaccharides eluted in the order of neutral, mono-, di-, tri- and tetra-sialylated species based on both charge, size and structure. Baseline separations were obtained with neutral oligosaccharide types but mixtures of high-mannose and complex types were poorly resolved. The high-mannose peaks were eliminated specifically from complex oligosaccharides by digesting with alpha-mannosidase. Treatment with beta-galactosidase, beta-N-acetylglucosaminidase and alpha-mannosidase resulted in a decrease of the oligosaccharide elution times corresponding to the number of sugar residues lost, the profile of changes was highly reproducible. In contrast, treatment with alpha-L-fucosidase, endo-beta-N-acetylglucosaminidase F and endo-beta-N-acetylglucosaminidase H resulted in an increase in their corresponding oligosaccharide retention times similar to the presence of an additional sugar residue. Conditions developed for separation of the reduced oligosaccharides and also a mixture of monosaccharide to oligosaccharide containing about 15 sugar residues within 30 min were useful in determining the effect of endo- and exo-glycosidases on porcine thyroglobulin oligosaccharides. Changes in elution time of the oligosaccharides following specific glycosidase digestions combined with methylation analysis provided a rapid and sensitive tool for confirmation of the carbohydrate primary structures present in thyroglobulin.
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PMID:Rapid characterization of asparagine-linked oligosaccharides isolated from glycoproteins using a carbohydrate analyzer. 199 74

A previous study has characterized the major 47 kDa anti-sticking factor (ASF-I) from goat cauda-epididymal plasma (Roy, N., and Majumder, G.C., Biochim. Biophys. Acta, 991:114-122, 1989). This study reports the purification and characterization of ASF-II, another anti-sticking factor from the goat epididymal plasma. ASF-II was purified to apparent homogeneity by using concanavalin A-agarose affinity chromatography, DEAE-cellulose chromatography, alumina gel adsorption, and isoelectric focussing techniques. It showed a single protein band by both non-denaturing and SDS-polyacrylamide gel electrophoresis. ASF-II showed a molecular weight of 36,000 and a sedimentation constant of 2.4S. ASF-II is largely stable to heat treatment and it is a specific glycoprotein having high affinity and specificity for its anti-sticking action. At saturating concentration (1 nM) it inhibited adhesion of nearly 50% of spermatozoa to the glass surface of the haemocytometer counting chamber. Both the protein and sugar parts of the factor are essential for the anti-sticking activity since it lost its activity completely when treated with trypsin, L-fucosidase, or mannosidase. ASF-II does not coat the glass surface and it binds to spermatozoa. Data are consistent with the view that ASF-II has not been derived from the larger ASF-I molecule due to its enzymic modifications. Both ASF-I and -II had no effect on sperm forward motility as evidenced by spectrophotometric motility assays, indicating thereby the suitability of the factors to improve the existing sperm motility assays by eliminating the possibility of cell-sticking artifacts.
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PMID:Characterization of anti-sticking factor-II from goat epididymal plasma. 209 69

A nonspecific inhibitor of macrophage migration was found in large quantities in the cell-free ascitic fluids from patients with ovarian tumours. MIF-like activity was assigned by the ability of various dilutions of ascitic fluids to inhibit migration of guinea pig macrophages from agarose droplets. The factor was purified in 3 subsequent steps including ion exchange chromatography, gel filtration, and isoelectric focusing. The data obtained indicate molecular heterogeneity according to net charge and molecular weights. The main MIF activity was found at about 45, 20, and 10 kD, respectively, and partially at less than 10 kD. Isoelectric focusing of the various MIF species revealed activity peaks in the pH range from 3.8 to 5.0. A further peak was detected at pH 6.0 in crude material. The factor was purified about 10 000-fold compared to the starting material. The action of OC-MIF was inhibited by L-fucose, and when target cells were incubated with alpha-L-fucosidase, they did not respond any longer to OC-MIF. Furthermore, purified MIF-like activity is a nondialyzable glycoprotein, sensitive to treatment with neuraminidase, chymotrypsin, trypsin and pronase, however, unaffected by incubation at 60 degrees C for 1 h. The physicochemical properties of OC-MIF activity studied are comparable to lymphocyte-derived conventional MIF.
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PMID:Inhibition of macrophage migration by a factor from ascites fluids of ovarian cancer patients. I. Biochemical characterization and purification. 351 10

MAT-B1 and MAT-C1 ascites sublines of the 13762 rat mammary adenocarcinoma both contain sialomucin as a major cell surface component and are resistant to cytolysis by normal rat spleen lymphocytes [3 +/- 2% (SD) and 0 +/- 1%, respectively]. Susceptibility to lysis did not increase following treatment of cells with neuraminidase, fucosidase, or alpha- or beta-galactosidase. Treatment with trypsin significantly increased the susceptibility of MAT-B1 (14 +/- 3%) but not MAT-C1 (5 +/- 2%). Following 1 month in culture, the sialomucin content of MAT-B1 cells dropped from 30% to 8% (determined by glucosamine labeling) and natural cell-mediated cytolysis increased to 16 +/- 4%, whereas the sialomucin content and susceptibility of MAT-C1 cells did not change. The results indicate that the relatively minor changes associated with removal of cell surface sialic acid or fucose residues do not result in increased susceptibility of the ascites cells to cytolysis. However, susceptibility of MAT-B1 cells to lysis by normal rat spleen lymphocytes was inversely correlated with the amount of major glycoprotein (r = -0.96).
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PMID:Cell surface sialomucin and resistance to natural cell-mediated cytotoxicity of rat mammary tumor ascites cells. 373 Nov 8

A study was made of the activity of 3 lysosomal glycosidases -beta-D-galactosidase (K. P. 3.2.1.23), alpha-L-fucosidase (K. P. 3.2.1.51), N-acetyl-beta-D-hexosoaminidase (K. P. 3.2.1.52) depending on the time after subcultivation and duration of the passage of human skin embryonal and postembryonal fibroblasts. It was established that changes in the specific activity of the enzymes should be calculated with reference to the cell rather than to protein whose amount might vary considerably. It was also found that for measuring the specific activity of enzymes, of great importance are the procedures of cell removal from the base layer (by mechanical scraping off or by trypsin solution) and the regimen of the homogenization of cell preparations.
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PMID:[Lysosomal glycosidase activity in cultured human fibroblasts]. 623 62

alpha-L-Fucosidase is a cell wall protein purified from pea (Pisum sativum) epicotyls. The alpha-L-fucosidase hydrolyzes terminal fucosyl residues from oligosaccharides of plant cell wall xyloglucan. alpha-L-Fucosidase may be an important factor in plant growth regulation, as it inactivates fucose-containing xyloglucan oligosaccharides that inhibit growth of pea stem segments. The amino acid sequences of the NH2-terminal region and one internal peptide were used to design redundant oligonucleotides that were utilized as primers in a polymerase chain reaction (PCR) with cDNA, generated from pea mRNA, as the template. A specific PCR amplification product containing 357 base pairs was isolated, cloned, and sequenced. The deduced amino acid sequence included the two peptides used to design the primers for PCR plus two other peptides obtained by proteinase digestion of alpha-L-fucosidase. No sequence homology to other alpha-L-fucosidases was apparent, although the NH2-terminal region is strongly homologous to Kunitz-type trypsin inhibitors. cDNA and genomic copies were isolated and sequenced. In pea, the gene is present in two or three copies. Its mRNA is present in roots, leaves, and elongating shoots. The spatial pattern of expression of the alpha-L-fucosidase was determined by in situ hybridization.
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PMID:Molecular cloning and pattern of expression of an alpha-L-fucosidase gene from pea seedlings. 755 5

The subunits of human liver alpha-L-fucosidase have been separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, excised, and subjected to peptide mapping after CNBr cleavage or trypsin digestion. The CNBr peptide maps of the glycosylated 56- and 51-kDa subunits were similar except that the larger subunit had several peptides with M(r)s shifted higher than those apparent for the smaller subunit. These M(r) differences were almost completely eliminated when CNBr peptide mapping was performed on the deglycosylated 48- and 45-kDa polypeptides, suggesting that the M(r) differences were due to carbohydrate differences. Minor differences not related to glycosylation were found in the CNBr peptide maps for the 48- and 45-kDa polypeptides including the presence of small amounts of three peptides in the larger polypeptide not found in the smaller polypeptide. Sequence analysis suggested that both the 48- and the 45-kDa polypeptides were blocked at their amino-termini but analysis of the largest CNBr peptide from each polypeptide indicated an identical 13-amino-acid sequence corresponding to residues 6 through 18 from the cDNA-deduced sequence of mature alpha-L-fucosidase. Tryptic peptide mapping indicated very similar HPLC peptide profiles for the deglycosylated 48- and 45-kDa polypeptides except for the presence of small amounts of six peaks present in the larger polypeptide which were not detected in the smaller polypeptide. The overall results provide the first evidence that the polypeptides of human liver fucosidase are very similar and probably encoded by the same gene. However, minor differences in the polypeptides exist, possibly due to normal allelic variation, alternative splicing, proteolytic processing, and/or posttranslational modifications other than those due to glycosylation.
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PMID:Peptide mapping of the subunits and deglycosylated polypeptides of human liver alpha-L-fucosidase. 803 Nov 25


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