Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.4 (trypsin)
 42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The blebbed surface morphology produced by trypsinisation of Chinese hamster ovary cells is subsequently reorganized to a microvillous topography, even in the continued presence of trypsin. Scanning and transmission electron microscope (SEM and TEM) observations of this transition showed the initial formation of a "crown' of densely clustered microvilli at one pole of the cell. At the periphery of this region the blebs coalesced to form ridges which subsequently extended over the entire cell surface. Long, and occasionally branched microvilli were generated from the ridges. Large numbers of membrane associated vesicles were also characteristic of these areas of surface reorganisation.
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PMID:Conversion of blebs to microvilli: cell surface reorganisation after trypsin. 732 17

The xylanase A (endo-1,4-beta-D-xylan xylanhydrolase) of the basidiomycete Schizophyllum commune was treated with the powerful carboxylate-modifying reagent 1-(4-azonia-4,4-dimethyl-pentyl)-3-ethylcarbodiimide iodide (EAC) in the presence of substrate. This treatment was followed by complete inactivation of the enzyme with [14c]EAC after the removal of excess reagent and protecting ligand. The inactivated enzyme was digested with endoproteinase Arg-C or trypsin, and peptides were separated and purified using reverse-phase high-performance liquid chromatography. Following sub-digestion of individual radioactive peptides with staphylococcal V8 protease and endoproteinase Lys-C, amino acid composition analysis and sequencing analysis revealed that the [14C]EAC label was bound exclusively to Glu87. Comparison of the primary sequences of related xylanase with that of xylanase A revealed that Glu87 is a highly conserved residue. Based on this similarity and the mechanism of carbodiimide action, Glu87 is proposed to act as the nucleophile in the catalytic mechanism of xylanase A. The possible environment of the putative catalytic glutamate residue was explored using hydrophobic-cluster analysis and secondary-structure prediction based on the primary sequence of xylanase.
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PMID:Identification of a glutamate residue at the active site of xylanase A from Schizophyllum commune. 790 49

The lamina basilaris of guinea pig cochlea was studied with SEM after trypsin treatment, and with TEM of resin sections and deep-etching replicas. The lamina consists of radial, evenly compacted filaments in the zona arcuata, and radial, discretely bundled filaments in the zona pectinata. In both zones, elementary filaments measured about 12 nm in thickness on the replica. The filaments formed more or less irregular passing bridges with each other and, eventually, a three-dimensional network which was continuous with the basement membrane under the supporting cells.
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PMID:Fine structure of the lamina basilaris of guinea pig cochlea. 829 28

Serratia fonticola CUV produces two isoenzymes (forms I and II) with beta-lactamase activity which were purified by a five-step procedure. The isoenzymes had identical kinetic parameters and isoelectric point (pI = 8.12). They were characterized by a specific activity towards benzylpenicillin of 1650 U/mg. The beta-lactamase hydrolyzed benzylpenicillin, amoxycillin, ureidopenicillins, first- and second-generation cephalosporins. Carboxypenicillins and isoxazolylpenicillins were hydrolyzed to a lesser extent. Towards cefotaxime and ceftriaxone (third-generation cephalosporins), the S. fonticola enzyme exhibited catalytic efficiencies much higher than those of MEN-1 and extended-spectrum TEM derivative beta-lactamases. The beta-lactamase from S. fonticola was markedly inhibited by beta-lactamase inhibitors such as clavulanic acid, sulbactam and tazobactam. The purified isoenzymes were digested by trypsin, endoproteinase Asp-N and chymotrypsin. Amino acid sequence determinations of the resulting peptides allowed the alignment of 267 amino acid residues (Swiss-Prot, accession number P 80545) for form I beta-lactamase. Form II is five residues shorter than form I at its N-terminus. From amino acid sequence comparisons, S. fonticola CUV beta-lactamase was found to share more than 69.3% identity with the chromosomally encoded beta-lactamases of Klebsiella oxytoca, Proteus vulgaris, Citrobacter diversus and the plasmid-mediated enzymes MEN-1 and Toho-1. Therefore, the oxyimino cephalosporin-hydrolyzing beta-lactamase of S. fonticola belongs to Ambler's class A. Contribution of the serine at ABL 237 in the broad-spectrum activity of these beta-lactamases is discussed.
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PMID:Characterization and amino acid sequence analysis of a new oxyimino cephalosporin-hydrolyzing class A beta-lactamase from Serratia fonticola CUV. 930 Aug 9

A casein kinase released from activated human platelets phosphorylates a number of plasma proteins extracellularly, and that activation of platelets in systemic lupus erythematosus patients parallels an increase in the phosphate content of plasma proteins, including C3. The present study was undertaken to characterize this platelet protein kinase and to further elucidate the effect(s) on C3 function of phosphorylation by platelet casein kinase. The phosphate content of human plasma C3 was increased from 0.15 to 0.60 mol phosphate/mol of C3 after platelet activation in whole blood or platelet-rich plasma. The platelet casein kinase was distinct from other casein kinases in terms of its dependence on cations, inhibition by specific protein kinase inhibitors, and immunological reactivity. C3 that had been phosphorylated with platelet casein kinase was tested for its susceptibility to cleavage by trypsin or the classical and alternative pathway convertases and its binding to EAC and IgG. Phosphorylation did not affect the cleavage of C3 into C3a and C3b, but the binding of fragments from phosphorylated C3 to EAC14oxy2 cells and to IgG in purified systems and in serum was increased by 1.6-4.5 times over that of unphosphorylated C3. A covariation was seen between the enhanced binding of C3 fragments to IgG after phosphorylation and an increased ratio of glycerol/glycine binding, from 2.0 for unphosphorylated C3 to 4.9 for phosphorylated C3. The present study suggests that an overall effect of phosphorylation of C3 by platelet casein kinase is to enhance the opsonization of immune complexes.
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PMID:Alterations in C3 activation and binding caused by phosphorylation by a casein kinase released from activated human platelets. 1035 96

The object of this study was to compare the extent of decellularization at each critical step of processing porcine skin to produce an acellular dermal matrix (ADM) for biomedical applications. The results demonstrated that the removal of epidermis using treatment with 0.25% trypsin for 18 h and 0.1% sodium dodecyl sulfate (SDS) for 12 h at room temperature was beneficial for the subsequent treatment to remove cells in the dermal structure. Lengthy incubation in 0.25% trypsin (12 h) and then 560 units/l Dispase (12 h) at 25 degrees C of small pieces of porcine skin from which the epidermis had been removed efficiently removed cells and cellular components from the skin. Histological examinations revealed that the epidermis, dermal fibroblasts, and epidermal appendages were completely removed by these treatments, and the basic dermal architecture of collagen bundles was that of a loose meshwork. Examinations by TEM showed that the characteristics of collagen fibers in the ADM were retained after complete removal of cells present under optimal conditions defined in this study. SDS-PAGE and size-exclusion HPLC revealed that collagen fibers in the ADM were mostly type I and showed two typical component peaks identified as oligomers and monomers, respectively.
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PMID:Process development of an acellular dermal matrix (ADM) for biomedical applications. 1475 54

The reactions of 6-methylidene penems 4-7 with beta-lactamases (TEM-1, SHV-1, Amp-C) were characterized by electrospray ionization mass spectrometry (ESI-MS). The kinetics of the reactions were monitored, demonstrating that only one penem molecule reacts to form an acyl-enzyme complex. For penem 5, the ESI-MS/MS spectrum of the hydrolysis product produced in the reaction was identical to the spectrum generated from a synthesized dihydro[1,4]thiazepine 10, confirming the rearrangement of the penem ring system to a seven-membered dihydro[1,4]thiazepine structure. Gas-phase ESI-MS/MS fragmentation data were rationalized due to tautomerization between imine and enamine substructures. ESI-MS/MS analysis of the T-6 trypsin-digested fragments of TEM-1 and SHV-1 demonstrated that the penems were only attached to Ser-70 of these class A beta-lactamases and that the penem ring structures were rearranged to seven-membered dihydro[1,4]thiazepines.
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PMID:Mechanism of inactivation of beta-lactamases by novel 6-methylidene penems elucidated using electrospray ionization mass spectrometry. 1521 94

In the current study, amine surface modified iron-oxide nanoparticles of 6 nm diameter without polymer coating were fabricated in an aqueous solution by organic acid modification as an adherent following chemical coprecipitation. Structure and the superparamagnetic property of magnetite nanoparticles were characterized by selected area electron diffraction (SAED) and superconducting quantum interference measurement device (SQUID). X-ray photoelectron spectrometer (XPS) and zeta potential measurements revealed cationic surface mostly decorated with terminal -NH(3)(+). This feature enables them to function as a magnetic carrier for nucleotides via electrostatic interaction. In addition, Fe(3)O(4)/trypsin conjugates with well-preserved functional activity was demonstrated. The nanoparticles displayed excellent in vitro biocompatibility. The NMR and the in vitro MRI measurements showed significantly reduced water proton relaxation times of both T(1) and T(2). Significantly reduced T(2) and T(2)*-weighted signal intensity were observed in a 1.5 T clinical MR imager. In vivo imaging contrast effect showed a fast and prolonged inverse contrast effect in the liver that lasted for more than 1 week. In addition, it was found that the spherical Fe(3)O(4) assembled as rod-like configuration through an aging process in aqueous solution at room temperature. Interestingly, TEM observation of the liver tissue revealed the rod-like shape but not the spherical-type nanoparticles being taken up by the Kupffer cells 120 h after tail vein infusion. Combining these results, we have demonstrated the potential applications of the newly synthesized magnetite nanoparticles in a broad spectrum of biomedical applications.
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PMID:Aqueous dispersions of magnetite nanoparticles with NH3+ surfaces for magnetic manipulations of biomolecules and MRI contrast agents. 1596 22

The aim of this study was to investigate the effect of trypsin digestion on removal of chondroitin sulfate proteoglycans (CS-PGs) in demineralized dentin, and subsequent dentin bonding. Bovine dentin fragments were demineralized, treated with or without trypsin, stained with cupromeronic blue, and observed under transmission electron microscopy. Demineralized sections with or without trypsin digestion were also subjected to immunohistochemical analysis with anti-chondroitin-4-sulfate (C4S) monoclonal antibody, 2-B-6. The presence of galactosamine and glucosamine in the trypsin digest was confirmed by amino acid analysis. Bond strength testing was performed on trypsin treated and control specimens where samples were either kept moist or dried and re-wet, then bonded. Bond strength significantly decreased after trypsin treatment (p < 0.05). TEM, immunohistochemical, and amino acid analyses demonstrated that trypsin digestion efficiently removed C4S-PGs from demineralized dentin matrix. This study indicates that the detrimental effects observed on dentin bonding by trypsinization may be due in part to the removal/cleavage of the C4S-PGs, and further underscore the importance of C4S-PGs on dentin bonding.
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PMID:Removal of dentin matrix proteoglycans by trypsin digestion and its effect on dentin bonding. 1791 48

Erythema Toxicum, a rash frequently present in the healthy newborn infant is an innate, immune response to the first commensal micro flora. Flushing and urtication are seen in this manifestation suggesting mast cell (MC) activation and MC derived mediator release. It has recently become evident that MCs participate in the protective, innate immune response against microbes also by secreting products toxic to pathogens such as cathelicidin peptide antibiotics. We hypothesized that MCs contribute to the process of inflammation in Erythema Toxicum and that skin MCs of human newborns express the cathelicidin peptide antibiotic LL-37. Skin sections were immunostained for MC tryptase. Double immunofluorescence was performed by staining LL-37 in combination with tryptase. We studied ultra structure of skin MCs with transmission (TEM) and immunoelectron microscopy (IEM). Seven infants with and six infants without the rash, as well as three adults were included. We found numerous tryptase-expressing MCs recruited around the hair follicles in the lesions of Erythema Toxicum. TEM analysis of MCs exhibited signs of degranulation in the lesion. Neither skin MCs from newborns nor adults did express LL-37 as judged by confocal and IEM. MCs participate in the inflammatory responses of Erythema Toxicum by taking an active part in the immune system of the hair follicle. However, their immunological activity is not linked to the expression of the cathelicidin antimicrobial peptide LL-37. A pivotal role of MCs in the innate, inflammatory response at the site of pathogen invasion during the critical time of perinatal colonization is suggested.
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PMID:Urticaria Neonatorum: accumulation of tryptase-expressing mast cells in the skin lesions of newborns with Erythema Toxicum. 1807 19


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