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)

In total, 86 enterococcal strains including representatives of most of the described species were tested for the ability to agglutinate human, sheep, and rabbit erythrocytes. Five strains did not react with any of the erythrocytes tested, and 81 (94.2%) strains agglutinated only rabbit erythrocytes. The hemagglutination titers ranged from 2 to 64. Loss of the hemagglutination activity was observed when rabbit erythrocytes were treated with trypsin or neuraminidase. Trypsin treatment of the bacterial suspensions also caused loss of the agglutination ability. On the other hand, heat treatment of bacterial suspensions increased the efficiency of the interactions, and higher titers were obtained. Assays for inhibition of hemagglutination were performed with alpha-D-fucose, alpha-D-galactose, beta-D-galactose, D-glucose, N-acetyl-galactosamine, N-acetyl-glucosamine, N-acetylneuraminic acid, N-acetylneuraminic acid-lactose, and fetuin. Only fetuin was able to inhibit the hemagglutination reactions. The results showed that hemagglutination properties are common to the different enterococcal species tested. They also suggest that enterococci possess hemagglutinins of proteic and non-proteic nature that are involved in the attachment to sialic acid-containing receptors on the surface of rabbit erythrocytes.
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PMID:Hemagglutination properties of Enterococcus. 776 54

In the accompanying study (Cho, M., and Cummings, R. D. (1995) J. Biol. Chem. 270, 5198-5206), we reported that Chinese hamster ovary (CHO) cells synthesize galectin-1. We have now used several approaches to define the subcellular location and biosynthesis of galectin-1 in these cells. Galectin-1 was present on the cell surface, as assessed by immunofluorescent staining with monospecific antibody to the protein. Quantitation of the surface-localized galectin-1 was achieved by metabolically radiolabeling cells with [35S]Met/Cys and measuring the amount of lectin (i) sensitive to trypsin, (ii) accessible to biotinylating reagents, and (iii) accessible to the haptenic disaccharide lactose. By all three procedures, approximately 1/2 of the radiolabeled galectin-1 associated with cells was shown to be on the cell surface with the remainder intracellular. The kinetics of externalization of galectin-1 was monitored by pulse-chase radiolabeling, and it was shown that cells secrete the protein with a t1/2 approximately 20 h. The cell surface form of galectin-1 in CHO cells was active and bound to surface glycoconjugates, but lectin accumulating in the culture media was inactive. Lectin synthesized by mutant Lec8 CHO cells, which are unable to galactosylate glycoproteins was not found on the surface and quantitatively accumulated in the media in an inactive form. Taken together, our results demonstrate that galectin-1 is quantitatively externalized by CHO cells and can associate with surface glycoconjugates where the lectin activity is stabilized.
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PMID:Galectin-1, a beta-galactoside-binding lectin in Chinese hamster ovary cells. II. Localization and biosynthesis. 789 Jun 31

Treatment of rabbit aortic endothelial cells, human umbilical vein endothelial cells, and human aortic endothelial cells for 4 hours with minimally oxidized low-density lipoprotein (MM-LDL) induced the adhesion of monocytes but not neutrophils or lymphocytes to these cells. This induction was blocked by inhibitors of glycoprotein synthesis (cycloheximide and tunicamycin), and binding was abolished by treatment of cells with low levels of trypsin, suggesting that the binding molecule(s) is a protein. There was no increase in binding of antibodies to E-selectin, vascular cell adhesion molecule-1 (VCAM-1), or intercellular adhesion molecule-1 (ICAM-1) after treatment of cells with MM-LDL. Treatment of endothelial cells with Fab fragments of antibody to monocyte chemotactic protein-1 or to fibronectin did not block monocyte binding. Several sugars (lactose-1-phosphate, maltose-1-phosphate, and N-acetylglucosamine) inhibited monocyte binding to cells treated with MM-LDL, but binding was not blocked by mannose-6-phosphate, fructose-6-phosphate, glucose-1-phosphate, or glucose-6-phosphate. EDTA or EGTA treatment inhibited binding, which was restored by adding either calcium or magnesium. We conclude that the binding of monocytes to endothelial cells induced by a 4-hour treatment with MM-LDL is caused by a binding molecule(s) other than E-selectin, VCAM-1, or ICAM-1 and that carbohydrate chains on the monocytes or the endothelium play a role in binding.
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PMID:Partial characterization of leukocyte binding molecules on endothelial cells induced by minimally oxidized LDL. 812 47

A galactoside-binding lectin (hL-31) containing a collagen-like sequence was identified in human tumor cells. It was found to be the homologue of the IgE-binding protein, the macrophage cell-surface Mac-2 antigen, and the murine CBP35, RL-29, and mL-34 lectins. Here we report on the expression in Escherichia coli and functional analysis of recombinant hL-31 (rhL-31). The rhL-31 was purified in one step through an asialofetuin affinity column. The rhL-31 was reactive to anti-lectin antibodies and retained its lactose-dependent hemagglutination of trypsin-treated glutaraldehyde-fixed rabbit erythrocytes. The rhL-31 elutes from an affinity column as a 31-kDa monomer and undergoes homodimerization at relatively high protein concentrations, comparable to those used to mediate hemagglutination. Electron microscopy showed that the rhL-31 appears as a Y-shaped structure. Lactoperoxidase-catalyzed iodination of murine tumor cell-surface proteins followed by collagenase treatment revealed that the lectin is probably a peripheral membrane protein whereby both the amino and the carboxy termini are exposed on the outer cell membrane. These results point to the membrane disposition and orientation of the lectin and suggest a mechanism for a structure-function relationship of lectin activity.
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PMID:Structure-function relationship of a recombinant human galactoside-binding protein. 847 70

Conditions affecting the adherence of clinical isolates of Cryptococcus neoformans to rat glial and lung cell cultures were studied. Adherence to glial cells was a time-dependent process that was affected by the yeast culture age and growth temperature. The most adherent yeasts were those from 48 h cultures grown at 37 degrees C. Formalin-treating the yeasts did not affect adherence but formalin-treating the glial monolayers prevented yeast binding. Treating the yeasts with trypsin reduced adherence to glial monolayers, indicating that the yeast adhesin had a trypsin-labile protein component. Certain carbohydrates inhibited cryptococcal adherence to glial and lung cells in a time and concentration-dependent manner. Of the carbohydrates tested, N-acetyl-D-glucosamine, sucrose, lactose, sorbitol and myo-inositol were the most inhibitory, while mannose, galactose and xylose were the least inhibitory. The results collectively indicated that the mechanisms of adherence of C. neoformans to lung cells were similar to those of glial cells and that both involved a protein-containing adhesin on the cryptococcal surface that was expressed only after growth at 37 degrees C. Carbohydrate receptors also appeared to be involved with these interactions.
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PMID:Conditions affecting the adherence of Cryptococcus neoformans to rat glial and lung cells in vitro. 848 58

A total of 259 Gram-negative Porphyromonas-like rods isolated from subgingival plaque samples of 16 family-owned dogs with naturally occurring periodontitis were characterized phenotypically by biochemical reactions, metabolic end products and enzymatic activities (API-ZYMTM, RoscoTM). Four distinct groups were found. Group A isolates (63) were asaccharolytic, lipase negative, trypsin positive and produced phenylacetic acid (PAA) from peptone-yeast extract glucose broth. Unlike P. gingivalis strains they were catalase positive. Group B isolates (42) differed from those of group A by a positive lipase reaction and from those of group D by failing to ferment sugars. Group C isolates (88) were asaccharolytic and did not produce PAA. They were alpha-fucosidase, N-acetyl-beta-glucosaminidase (beta-NAG) and trypsin negative, resembling P. endodontalis, but unlike human isolates, they were catalase positive. Subgroup C.1 isolates (6) differed from those of parent group C by producing minor amounts of PAA, and subgroup C.2 isolates (12) were beta-NAG positive. Group D isolates (46) were weakly fermentative, lipase, catalase and trypsin positive, and produced PAA. They resembled the B (P.) salivosus type strain which, in our hands, fermented weakly glucose, lactose and mannose. Two isolates could not be assigned to any of the previous groups.
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PMID:Porphyromonas-like gram-negative rods in naturally occurring periodontitis in dogs. 851 57

Galactosyltransferase and UDP-[3H]galactose are commonly used to identify O-linked N-acetylglucosamine (O-GlcNAc)-bearing proteins and peptides. In this report we show that immobilized Ricinus communis agglutinin I (RCA I) specifically binds in vitro galactosylated O-GlcNAc-bearing peptides, facilitating their selective isolation from complex mixtures. First, the peptide YSDSPSTST was O-GlcNAc glycosylated, galactosylated, and sialylated. Of these three glycoforms, only the one with a terminal galactose interacted with the lectin. Next, RCA I was used to isolate glycopeptides from the O-GlcNAc-bearing basic phosphoprotein (BPP) of human cytomegalovirus. BPP was overexpressed using baculovirus, [3H]galactosylated, digested with trypsin, and fractionated on RCA I. Peptides that were not galactosylated passed through the column, whereas the majority of the radiolabeled glycopeptides interacted weakly with the lectin and did not require lactose or elution. These radiolabeled peptides eluted as a broad peak with the leading edge being characterized by more hydrophobic glycopeptides and the lagging edge by less hydrophobic peptides, suggesting that the polypeptide backbone may influence the interaction with the lectin. Lactose was required to elute the remaining radiolabeled peptides, suggesting that these peptides are multiply glycosylated. The weakly interacting glycopeptides were analyzed directly by liquid chromatography/electrospray-mass spectrometry (LC/ES-MS). Glycopeptides corresponding to both of the major sites of glycosylation of BPP were identified. Thus, RCA I greatly facilitates the selective isolation of in vitro galactosylated O-GlcNAc glycopeptides from complex mixtures and substantially reduces the purification required for subsequent site-mapping by gas-phase sequencing and/or LC/ES-MS.
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PMID:Specific isolation of O-linked N-acetylglucosamine glycopeptides from complex mixtures. 857 67

Bacterial surface hydrophobicity (SH) plays a role in adhesion of bacteria to host surfaces and ingestion by phagocytic cells. Streptococcus dysgalactiae (n = 60) isolated from bovine intramammary infections were examined for expression of SH after growth in Todd-Hewitt broth (THB) and THB supplemented with skim milk, whey, lactose, and casein. Strains were significantly more hydrophobic after growth in THB and THB plus whey and more hydrophilic after growth in THB plus skim milk. Both trypsin and proteinase K abolished SH in three strains tested. Mild pepsin treatment had little effect on SH, while heat treatment at 70 degrees or 80 degrees C abolished SH in two strains tested. A hydrophilic strain of S. dysgalactiae did not adhere as well to bovine mammary epithelial cells as a hydrophobic strain. Trypsin treatment significantly reduced adherence of a hydrophobic strain of S. dysgalactiae to epithelial cells while adherence of a hydrophilic strain remained unaltered. A hydrophilic strain of S. dysgalactiae was significantly more resistant to phagocytosis by bovine mammary gland macrophages than a hydrophobic strain. Differences in expression of SH may play an important role in determining the ability of S. dysgalactiae to establish successfully within the mammary gland.
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PMID:Influence of Streptococcus dysgalactiae surface hydrophobicity on adherence to mammary epithelial cells and phagocytosis by mammary macrophages. 877 7

To investigate what factors lead to rapid postnatal tissue growth and functional maturation in the newborn intestine, we compared intestinal tissue mass and digestive enzyme activities between newborn unsuckled piglets and piglets bottle fed for 3 days with either 5% lactose solution, intact porcine colostrum or trypsinized porcine colostrum. Bottle feeding of colostrum or trypsinized colostrum, but not lactose solution, led to a significant increase in the weight and length of the small intestine (p < 0.01) and a significant increase in the mucosal weight of the large intestine (p < 0.05). The mucosal protein content in the small and large intestine and the mucosal DNA content in the large intestine increased significantly following 3 days of bottle feeding of porcine colostrum or trypsinized colostrum. The total mucosal DNA contents in the small intestine of piglets fed colostrum or trypsinized colostrum were, respectively, 39 and 64% greater than that in the newborn unsuckled piglets. Intestinal digestive enzymes showed a differential response to the dietary treatment. Bottle feeding of intact porcine colostrum, but not trypsinized porcine colostrum led to a significant increase in lactase- and alkaline phosphatase-specific activities in the small intestine, while bottle feeding of lactose solution led to a significant decrease in the specific activity of lactase. In contrast, the specific activity of maltase in the small intestine increased significantly with age irrespective of dietary treatment. These results indicate that genetic and dietary factors are involved in regulating postnatal intestinal development, and porcine colostrum contains a trypsin-labile component which can increase lactase and alkaline phosphatase activities in the newborn intestine.
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PMID:Effects of colostrum feeding on intestinal development in newborn pigs. 900 95

This paper describes a systematic procedure for introducing protease-sensitive sites into bacterial integral membrane proteins. Such sites should make it possible to monitor the subcellular localization of individual domains of a topologically complex protein. Escherichia coli lac permease was used as a model. Site-directed mutagenesis, targeted to a particular periplasmic domain, was used to generate insertion derivatives containing a lysine residue in different sequence contexts. Individual mutants were then screened for lactose transport activity and efficient cleavage by trypsin. To facilitate this screen, the mutagenesis was carried out using a gene fusion encoding an easily detected, bifunctional lac permease-galactosidase hybrid. Insertions were identified in the fourth and sixth periplasmic domains (P4 and P6) that were efficiently cleaved in both the hybrid protein and in unfused lac permease. One of the P6 insertion mutants exhibited lactose transport specific activity near that of the wild-type and was shown by sequence analysis to be cleaved at the expected site in the inserted sequence. As part of this analysis, we determined the range of cellular concentrations of lac permease over which lactose uptake was linear. The activity showed a plateau at a relatively low concentration corresponding to approximately five times the wild-type level.
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PMID:Engineering trypsin-sensitive sites in a membrane transport protein. 927 86


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