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)

Lactase-phlorizin hydrolase (LPH) (EC 3.2.1.23/62) is a major intestinal microvillar membrane glycoprotein that digests lactose, the main carbohydrate of milk. To investigate structure/function relationships of LPH and to assess the impact of intracellular processing on the function of LPH and on its transport to the cell surface, we have expressed a full-length cDNA encoding LPH in mammalian COS-1 cells. Analysis of the expressed protein by immunoprecipitation with monoclonal anti-LPH antibodies and treatments with endo-beta-N-acetylglucosaminidase H and sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed two polypeptides with apparent molecular masses of 215 and 230 kDa, representing the mannose-rich (pro-LPHh) and complex (pro-LPHc) glycosylated forms of the precursor. By contrast to pro-LPH in human enterocytes, the expressed pro-LPH in COS-1 cells does not undergo intracellular proteolytic cleavage to generate a form similar to the mature enzyme of the brush-border membrane. Intracellular cleavage, however, is not essential for the molecule to acquire its enzymatic activity since pro-LPH in COS-1 cells is enzymatically as active as LPH isolated from intestinal brush-border membranes. Indirect immunofluorescent staining of transfected cells demonstrated that pro-LPH is expressed at the cell surface. This was further corroborated by the sensitivity of the complex glycosylated form (pro-LPHc) to trypsin in the medium. Our results provide the first conclusive evidence that pro-LPH is an enzymatically active molecule and that the intracellular proteolysis of pro-LPH is not essential for the generation of transport-competent forms of LPH.
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PMID:Expression of a full-length cDNA coding for human intestinal lactase-phlorizin hydrolase reveals an uncleaved, enzymatically active, and transport-competent protein. 190 19

Intergeneric bacterial adherence is responsible for the complexity of the microbiota in human dental plaque and is believed to enable some extraneous bacteria to initially colonize the human oral cavity. Some current evidence indicates that Streptococcus sanguis, an early colonizer of teeth, enhances subsequent colonization by Porphyromonas (Bacteroides) gingivalis, a bacterium associated with advanced adult periodontitis. In this study, selected strains of P. gingivalis and S. sanguis were tested for their adherence activities in vitro. A differential filtration assay was devised in which one member of the test pair was radiolabeled. Heterogeneous aggregates that formed in mixed suspensions were collected on polycarbonate filters (8-microns pore size) and were washed free of individual bacteria and small homologous clumps. P. gingivalis 381, W50, JKG7, and 33277 adhered to S. sanguis G9B, M5, Challis 6, and 38. P. gingivalis A7A1-28 did not adhere well to S. sanguis under these conditions. More precise measurements of intergeneric adherence were obtained with an alternative assay with radiolabeled P. gingivalis and an artificial dental plaque composed of S. sanguis coupled to cyanogen bromide-activated agarose beads. CNBr-agarose was selected as the supporting matrix for the plaque because it was uniformly and permanently coated with S. sanguis and because P. gingivalis had negligible adherence activity for streptococcus-free beads. P. gingivalis W50 grown to the early stationary phase adhered to S. sanguis-coated beads in higher numbers than either midlogarithmic- or late-stationary-phase cells. Intergeneric adherence was not inhibited or reversed by the presence of lactose or other monosaccharides or disaccharides. Pretreatment of either bacterium with trypsin or proteinase K reduced subsequent adherence by 86 to 100%. Neuraminidase treatment of P. gingivalis caused 98% reduction of adherence, whereas similar treatment of S. sanguis caused only a 2% loss. Preincubation of P. gingivalis at 60 degrees C for 30 min decreased subsequent adherence to S. sanguis-coated beads by 94%. Adherence was reduced by 96% when bacteria were assayed while suspended in human whole saliva or when pretreated with saliva and subsequently assayed in buffer. The concentration of whole human saliva required to inhibit 50% adherence in this assay was 23 micrograms per ml (1:200 dilution). Suspension of the bacteria in normal rabbit serum resulted in 94% inhibition of adherence. These data indicate that saliva and serum may be important host defense factors for controlling Porphyromonas-Streptococcus adherence.
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PMID:Adherence of Porphyromonas (Bacteroides) gingivalis to Streptococcus sanguis in vitro. 198 21

Oral isolates of Streptococcus milleri were examined for their ability to coaggregate with actinomyces. Of the 68 S. milleri strains tested, including 3 reference strains, 40 strains coaggregated with Actinomyces naeslundii WVU45 (actinomyces coaggregation group B) and 36 strains coaggregated with Actinomyces viscosus T14V (actinomyces coaggregation group A). All S. milleri strains of serotypes b (4 strains), e (2 strains), and f (24 strains) coaggregated with both of the actinomyces. The coaggregation reactions between the S. milleri cells and A. naeslundii WVU45 cells were optimal at about pH 7.0 and were Ca2+ or Mg2+ dependent, but they were not inhibited by the presence of simple sugars or amino sugars, including lactose (up to 0.5 M). Treatment of the S. milleri cells with heat (100 degrees C, 3 min) or proteases (trypsin, 1.0 mg/ml; pronase, 0.25 mg/ml; 37 degrees C; 3 h) and of the actinomyces cells with periodate (0.01 M, 4 degrees C, 16 h) destroyed their coaggregating abilities. The coaggregations between cells of the S. milleri strains, we well as cells of the Streptococcus sanguis H1 (reference strain for streptococcus coaggregation group 2) and the actinomyces strains (WVU45 and T14V), were inhibited by AFH1 (a carbohydrate receptor on T14V cells for a lectin on H1 cells). These interactions were also inhibited by anti-AFH1 immunoglobulin G (IgG) and by anti-b, anti-e, and anti-f S. milleri IgG or anti-f IgG Fab fragments. These results suggest that S. milleri, at least strains of serotypes b, e, and f, belongs to streptococcus coaggregation group 2.
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PMID:Cellular coaggregation of oral Streptococcus milleri with actinomyces. 229 47

Campylobacter pylori is the causative agent of gastritis and possibly of peptic and duodenal ulcers in adults. Histological observations show C. pylori attached to gastric epithelium as well as in the mucus layer of the stomach. We found that clinical isolates of C. pylori possess a cell-bound hemagglutinin detectable with human erythrocytes (all phenotypes tested) and those of a variety of animal species. The C. pylori hemagglutinin is antigenic, heat sensitive, and destroyed by pronase and papain but resistant to pepsin and trypsin. The hemagglutinin has fibrillar morphology; C. pylori-erythrocyte interaction displays very intimate contact, which is typical of fibrillae-mediated attachment. Fibrillae were removed from C. pylori by solubilization with N-octylglucose. After partial purification and removal of N-octylglucose by dialysis, the protein reaggregated, with the assembly of fibrillar structures. Hemagglutination inhibition was observed with the sialoproteins fetuin, alpha 2-macroglobulin, and glycophorin A but not with asialofetuin or asialoglycophorin A. The erythrocyte receptor was more sensitive to destruction by a neuraminidase specific for the N-acetylneuraminyl-alpha(2-3)-galactopyranosyl [NeuAc(2-3)Gal] sequence than one specific for NeuAc(2-6)Gal. Hemagglutination-inhibition assays with N-acetylneuraminyl-alpha(2-3)-lactose [NeuAc(2-3)-lactose] and NeuAc(2-6)-lactose confirmed that the C. pylori hemagglutinin preferentially binds to the NeuAc(2-3)Gal isomer of NeuAc-lactose. Based upon the above-described properties of the C. pylori fibrillar hemagglutinin, we conclude that this antigen should be designated as a putative colonization factor antigen.
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PMID:N-acetylneuraminyllactose-binding fibrillar hemagglutinin of Campylobacter pylori: a putative colonization factor antigen. 245 65

Bluetongue virus (BTV) was shown to agglutinate human, ovine and porcine erythrocytes. Removal of neuraminic acid (NA) from erythrocytes by Vibrio cholerae neuraminidase prevented their agglutination. Haemagglutination was also inhibited by N-acetyl neuraminic acid (NANA), N-glycol neuraminic acid (NGNA) and N-acetyl neuramin-lactose. The ability of BTV to agglutinate trypsin-treated human erythrocytes, which lack the amino-terminal domain and the single N-linked oligosaccharide of glycophorin A, suggests that the virus bound to human erythrocytes via NANA-containing, O-linked oligosaccharides. Glycoproteins with NA-containing oligosaccharides of known structure such as mucin, fetuin, alpha 1-acid glycoprotein, ovomucoid and ovine, porcine, human and equine glycophorin were examined for their ability to inhibit BTV-mediated agglutination of human, ovine and porcine erythrocytes. All glycoproteins containing NANA- or NGNA alpha 2-6GalNAc were capable of inhibiting the agglutination of human and porcine erythrocytes. Treatment of human erythrocytes with Newcastle disease virus neuraminidase and of porcine erythrocytes with Clostridium perfringens neuraminidase to cleave preferentially the NANA- and NGNA alpha 2-3Gal linkages respectively, were shown to have little effect on the ability of the erythrocytes to be agglutinated by BTV. The results suggested that BTV binds to NANA- and NGNA alpha 2-6GalNAc residues in the O-linked oligosaccharides of human and porcine glycophorins respectively and indicated the presence of different binding sites on the virus for erythrocytes from other species.
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PMID:The site of bluetongue virus attachment to glycophorins from a number of animal erythrocytes. 255 61

The development of the human fetal gastrointestinal tract takes place early during gestation. The pancreas although developed by morphological means at the 16th week of gestation excretes its exocrine enzymes later at the 24th week of gestation except for amylase which reaches its full activity 6 months after birth. Trypsinogen secreted at the 24th week is activated into trypsin by enterokinase at the 26th week of gestation whereas lipase and colipase are secreted from the 24th week. The small intestine starts developing at the 10th week morphologically and functionally. At the same time when villi and crypts start to develop at the 11th to 12th week the first enzyme activities can be detected, i.e. sucrase-isomaltase, maltase-glucoamylase and lactase. Also peptidases and lysosomal hydrolases are measured at this age. With the exception of lactase, intestinal enzymes reach sufficient activities at the 25th week of gestation. Lactase activity remains low until the 32nd-34th week. For the digestion and absorption of lipids, protein and carbohydrates the gastrointestinal tract of premature infants under 1500 g in rather well equipped. Lipids are hydrolysed by the mutual action of breast milk lipase, lingual lipase, gastric lipase and pancreatic lipase. The carbohydrates lactose and oligosaccharides as supplements to breast milk are hydrolysed by lactase, sucrase-isomaltase and maltase-glucoamylase. Breast milk proteins and cows milk hydrolysates are digested by pancreatic proteases into oligopeptides which can be hydrolysed within the lumen by brush border peptidases and be absorbed. Peptides also can actively be transported through the microvillus membrane and be hydrolyzed by intracellular peptidases.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:[Nutrition of premature infants below 1,500 g: enteral prerequisites]. 309 34

Oligosaccharide recognition by the 13 kDa soluble lectin from bovine heart muscle has been investigated by inhibition of binding of the 125I-labelled lectin to trypsin-treated rabbit erythrocytes. The results indicate that the Type 1 (Gal beta 1-3GlcNAc) and the Type 2 (Gal beta 1-4GlcNAc) backbone structures are the basic recognition units, and that the blood-group-H structure, the blood-group-B structure, the 'B-like' structure [afucosyl-(blood group B)] and the alpha 2-3 sialylated analogues of the backbone structures can also be accommodated and hence are candidate receptor structures for the lectin. A comparison of available inhibition data on six other soluble beta-galactoside-binding lectins (three from human lung and three from rat lung) has shown some common features among these and the bovine lectin, e.g. in general a stronger reaction with N-acetyl-lactosamine than with lactose, and a lack of reaction with 3-fucosyl-lactose and 6-sialyl-lactose. However, there are distinctive features among the lectins, e.g. differences in relative reactions with the blood-group-A structure, and no two of the lectins appear to be identical in their fine specificities.
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PMID:Further studies of oligosaccharide recognition by the soluble 13 kDa lectin of bovine heart muscle. Ability to accommodate the blood-group-H and -B-related sequences. 342 6

The amino acid sequence of thiogalactoside transacetylase, a dimer, has been determined. The monomer contains 202 amino acid residues in a single polypeptide chain and has a molecular weight of 22,671. The analysis was carried out by treatment of the carboxymethylated protein with cyanogen bromide and with trypsin. All seven cyanogen bromide peptides were isolated in pure form and were ordered by peptides isolated from tryptic digests. The sequence analysis was aided by determination of the DNA sequence of the lacA gene. The amino terminus of the protein is heterogenous because the initiator methionine is only partially cleaved. Another rather unusual feature of this cytoplasmic protein is a very hydrophobic segment in the center portion of the chain. Comparison of the amino acid sequence of thiogalactoside transacetylase to those of the lac repressor, beta-galactosidase, and lactose permease did not reveal any marked similarities. Therefore, there is no obvious evolutionary relatedness among proteins of the Lactose Operon.
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PMID:The amino acid sequence of thiogalactoside transacetylase of Escherichia coli. 392 33

Neonatal maturation of alveolar structure in rat lung temporally coincides with the peak activity of a beta-galactoside-specific endogenous lung lectin of the type that has been linked to developmental processes in other tissues. To learn more about the lung lectin we examined four cell types for their ability to synthesize and bind the lectin. Cultured lung fibroblasts and pulmonary artery endothelial cells synthesized and bound the lectin, alveolar macrophages bound but did not synthesize it, and alveolar epithelial type II cells neither bound nor synthesized it. Pulmonary macrophages made a different lactose-binding protein that did not agglutinate or bind trypsin-treated red blood cells, a property of lung lectin that is the means of its assay.
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PMID:Synthesis and binding of lactose-specific lectin by isolated lung cells. 397 75

The necessity of developing a quick, sensitive, and reliable test for Salmonella in nonfat dry milk (NDM) is evident from the recent tracing of Salmonella outbreaks to this product. Normally, coagulation of casein occurs when assaying NDM under regular cultural conditions, raising the possibility of trapped bacteria. After 20 hr of incubation of NDM in preenrichment lactose broth, enrichment was achieved by using Selenite-Cystine Broth. Smears from the enrichment broth were examined by the fluorescent-antibody technique (FAT) with a commercially available polyvalent O globulin conjugated with fluorescein. Standard cultural methods (SCM) were performed for comparison with FAT. Sensitivity of FAT was definitely improved by the use of trypsin. Casein coagulation of NDM can be avoided by addition of trypsin to samples during initial preenrichment in lactose broth. Samples containing approximately one Salmonella per 10 g were easily detected by FAT with the use of trypsin-treated samples. The method required only 42 hr to complete. Additionally, the use of trypsin enhanced recovery of Salmonella by use of SCM, as evidenced by alteration in the observed coliform to Salmonella ratios.
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PMID:Increased sensitivity of immunofluorescent assay for Salmonella in nonfat dry milk. 490 97


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