Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.24.27 (thermolysin)
 1,894 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An inhibitor of neutral subtilopeptidase [EC 3.4.24.4] was purified from porcine serum by salting out with (NH4)2SO4, chromatography on anion exchange sephadex, gel filtration with Sepharose 6B, and isoelectric focusing. The preparation was homogeneous by electrophoretic and ultracentrifugal criteria, and was shown to be a glycoprotein with a molecular weight of 740,000. It inhibited the caseinolytic activities of thermolysin, subtilisin, trypsin [EC 3.4.21.4], and alpha-chymotrypsin [EC 3.4.21.1] as well as that of neutral subtilopeptidase by an equimolar binding to those proteolytic enzymes. SDS-polyacrylamide gel electrophoresis after reduction with beta-mercaptoethanol indicated that the inhibitor was made up of four subunit monomers having a molecular weight of 190,000. From comparisons of its physiocochemical and inhibitory properties with those of well-investigated plasma proteins, the inhibitor was identified as alpha2-macroglobulin. On treatment of the inhibitor with neutral subtilopeptidase, a protein with a molecular weight of 95,000 appeared after treatment with SDS and beta-mercaptoethanol, suggesting that a peptide bond susceptible to the enzyme exists near the mid-point of the subunit chains.
 ...
PMID:A neutral subtilopeptidase inhibitor from porcine serum some evidence for alpha2-macroglobulin. 5 60

The proteolytic enzyme, thermolysin, degraded the external segment of the membrane glycoprotein of intact vesicular stomatitis (VS) virions but left behind a small nonglycosylated fragment, presumably embedded in the virion membrane. Other proteases generated membrane-associated glycoprotein fragments differing somewhat in molecular weight. The thermolysin-resistant, virion-associated fragment, which can be selectively solubilized by either Triton X-100 or chloroform/methanol, has a molecular weight of 5,200. Amino acid analysis of the glycoprotein fragment reveals a preponderance of hydrophobic amino acids (64% of the residues); the amino-terminal amino acid is alanine as determined by dansylation. Cyanogen bromide digestion of the tail fragment generated two peptides, confirming the presence of one methionine residue per thermolysin-resistant glycoprotein fragment. The secondary structure of this glycoprotein tail peptide is maintained by at least one disulfide bridge. Thermolysin treatment is isolated VS viral glycoprotein in the presence of Triton X-100 also generated a hydrophobic peptide fragment which is very similar to the virion-associated glycoprotein fragment. The amino acid terminus of intact glycoprotein was also found to be alanine as was its dansylated Triton-micellar fragment that resisted thermolytic degradation; this finding suggests that the amino-terminal end of the VS viral glycoprotein is embedded in the virion membrane. These results suggest that the VS viral glycoprotein is an amphipathic molecule, the hydrophilic portion of which contains all the carbohydrate and a lipophilic tail segment which forms lipid or detergent micelles, thus rendering it resistant to proteolysis.
 ...
PMID:Association of vesicular stomatitis virus glycoprotein with virion membrane: characterization of the lipophilic tail fragment. 16

We have observed that treatment of rabbit synovial fibroblasts with proteolytic enzymes can induce secretion of collagenase (EC 3.4.24.7) and plasminogen activator (EC 3.4.21.-). Cells treated for 2-24 hr with plasmin, trypsin, chymotrypsin, pancreatic elastase, papain, bromelain, thermolysin, or alpha-protease but not with thrombin or neuraminidase secreted detectable amounts of collagenase within 16-48 hr. Treatment of fibroblasts with trypsin also induced secretion of plasminogen activator. Proteases initiated secretion of collagenase (up to 20 units per 10(6) cells per 24 hr) only when treatment produced decreased cell adhesion. Collagenase production did not depend on continued presence of proteolytic activity or on subsequent cell adhesion, spreading, or proliferation. Routine subculturing with crude trypsin also induced collagenase secretion by cells. Secretion of collagenase was prevented and normal spreading was obtained if the trypsinized cells were placed into medium containing fetal calf serum. Soybean trypsin inhibitor, alpha(1)-antitrypsin, bovine serum albumin, collagen, and fibronectin did not inhibit collagenase production. Although proteases that induced collagenase secretion also removed surface glycoprotein, the kinetics of induction of cell protease secretion were different from those for removal of fibronectin. Physiological inducers of secretion of collagenase and plasminogen activator by cells have not been identified. These results suggest that extracellular proteases in conjunction with plasma proteins may govern protease secretion by cells.
 ...
PMID:Proteases induce secretion of collagenase and plasminogen activator by fibroblasts. 20 72

The glycoprotein of vesicular stomatitis (VS) virus was selectively liberated from the virion membrane by the dialyzable nonionic detergent, beta-D-octylglucoside. The isolated viral glycoprotein could be rendered virtually free of phospholipid and detergent, under which conditions it formed tail-to-tail glycoprotein micelles in the form of rosettes. When mixtures of viral glycoprotein and egg lecithin were dialyzed free of octylglucoside, glycoprotein vesicles formed spontaneously with spikes protruding in the same external orientation as the VS virion membrane. The glycoprotein vesicles exhibited increased and uniform buoyant density, indicating relative homogeneity in the proportion of glycoprotein and phosphatidylcholine in each glycoprotein liposome. Evidence for similar insertion and orientation of VS viral glycoprotein in both phosphatidylcholine vesicles and virion membrane was substantiated by the finding that proteolytic digestion with thermolysin gave rise to hydrophobic glycoprotein tail fragments in vesicle or virion membranes that migrated identically in polyacrylamide gels.
 ...
PMID:Reconstitution into liposomes of the glycoprotein of vesicular stomatitis virus by detergent dialysis. 22 Feb 38

Selective binding of lipid to glycoprotein was detected when [3H]palmitate-labeled Sindbis virus particles or viral-infected cells were disrupted by heating with sodium dodecyl sulfate, and glycoproteins were isolated by electrophoresis in sodium dodecyl sulfate/10% polyacrylamide gels. The smaller glycoprotein (E2) retained 2 to 3 times more labeled lipid than did the larger EI glycoprotein, and the cell-associated glycoprotein precursor (PE2) bound even less lipid. No lipid was associated with the nonglycosylated glycoproteins that accumulated in infected cells treated with tunicamycin. The labeled lipid remained bound to the glycoproteins after exhaustive extraction with chloroform/methanol of virus particles, infected-cell extracts, or isolated glycoproteins, but it could be extracted by chloroform/methanol after treating glycoproteins with mild alkali. Analysis by gas/liquid chromatography showed that 60% of the label was in palmitate and the balance of label was distributed between oleate and stearate. There were approximately 2 mol of fatty acid bound per mol of E1 glycoprotein. Proteolysis of the fatty acid-labeled glycoprotein with pepsin, thermolysin, and Pronase degraded the polypeptide to fragments that retained the fatty acids in an alkali-labile state. These data suggest that a covalent attachment of fatty acid may occur during maturation of the viral glycoproteins.
 ...
PMID:Evidence for covalent attachment of fatty acids to Sindbis virus glycoproteins. 28 8

The cell-free haemolymph of the mollusc Octopus vulgaris inhibited the proteolytic activity of the thermolysin against the high-molecular-mass substrate hide powder azure. The purified inhibitor was a glycoprotein composed of two identical 180 kDa disulphide-linked subunits. In addition to the inhibition of the metalloproteinase thermolysin, the protein inhibited the serine proteinases human neutrophil elastase, pig pancreatic elastase, bovine chymotrypsin, bovine trypsin and the cysteine proteinase papain. A fraction of the proteinase-inhibitor complex resisted dissociation after denaturation indicating that some of the proteinase molecules became covalently bound. The nucleophile beta-aminopropionitrile decreased the covalent binding of proteinases to the Octopus vulgaris protein, suggesting that this interaction is mediated by an internal thiol ester; the reactivity and the amino acid sequence flanking the reactive residues of the putative thiol ester were consistent with this hypothesis. Bound trypsin remained active against the low-molecular-mass chromatogenic substrate H-D-Pro-Phe-Arg p-nitroanilide and was protected from inhibition by active-site-directed protein inhibitors of trypsin; however, the bound trypsin was readily inhibited by small synthetic inhibitors. This indicates that the inhibition of proteinases is accomplished by steric hindrance. The proteinase-inhibitory activity of this protein is characteristic of inhibition by mammalian alpha-macroglobulins and the presence of a putative thiol ester suggests that the Octopus vulgaris proteinase inhibitor is a homologue of human alpha 2-macroglobulin.
 ...
PMID:Purification and characterization of an alpha-macroglobulin proteinase inhibitor from the mollusc Octopus vulgaris. 137 44

Characteristic properties of the antigens recognized by sperm-immobilizing monoclonal antibodies (SI-mAbs) from different sources were compared by ELISA competitive inhibition assay, Western blot analysis, chromatographic analysis, and enzymatic digestion studies. Among 9 SI-mAbs, human mAb H6-3C4 and three mouse mAbs--2C6, 2B6, and 2E5--also possessed strong sperm-agglutinating activity. Binding of human mAb H6-3C4 to sperm was strongly inhibited by the three mouse mAbs (2C6, 2B6, and 2E5), but not by the rat or the other four mouse mAbs. SDS-PAGE revealed that mAb H6-3C4 and three mouse mAbs recognized the same antigen molecules of 15-25 kDa present in both sperm extracts and seminal plasma. Chemical treatments with trifluoromethanesulfonic acid and sodium metaperiodate destroyed the antigen determinants recognized by the above four mAbs, as detected by both ELISA and antibody absorption tests. Western blot analysis revealed that the antigens were susceptible to treatments with papain, proteinase K, and N-glycanase, but resistant to trypsin, V8 protease, and thermolysin. These results indicate that one of the major antigens recognized by mAbs with sperm-immobilizing action may be a sperm membrane-associated glycoprotein of 15-25 kDa and the epitope may involve N-linked oligosaccharides.
 ...
PMID:Comparative studies of the antigens recognized by sperm-immobilizing monoclonal antibodies. 161 9

Echinonectin (EN) is a 230-kDa extracellular matrix glycoprotein found in the hyaline layer of sea urchin embryos. Dissociated embryonic cells attached strongly to EN-coated microtiter wells in a centrifugal-based in vitro adhesion assay, suggesting that EN is one of the hyaline layer proteins to which cells adhere in vivo (Alliegro et al., 1988). The present study examines the molecular properties of that adhesion using monoclonal antibodies as probes to block cell attachment, and also demonstrates that EN possesses lectin activity. EN binds tenaciously to agarose-based chromatography resins, such as Sepharose. The sugar-binding activity is associated with the polypeptide component of EN, and not with the carbohydrate moiety. Binding is inhibited with galactose and fucoidan, but not with glucose or locust bean gum. Although functional sites both for polysaccharide binding and for cell attachment are present on each subunit of the EN molecule, the sites appear to be functionally distinct because galactose and fucoidan are completely without effect on cell attachment in vitro. Proteolytic digestion of EN yields a highly limited set of immunoreactive peptides. Digestion with trypsin yields a 20-kDa fragment, chymotrypsin, a doublet at 20 kDa, and 20- and 23-kDa fragments with thermolysin. McAb's directed against these peptides block cell adhesion in vitro, suggesting that they possess the cell attachment domain of EN. This is supported by the observations that trypsin-digested EN is an effective substrate in adhesion assays and that adhesion to the tryptic fragments is also blocked by McAb's to the 20-kDa domain.
 ...
PMID:In vitro biological activities of echinonectin. 232 45

Chlamydomonas flagellar sexual agglutinins are responsible for the adhesion of opposite mating-type (plus and minus) gametes during the first stages of mating. Purification and partial characterization of the plus agglutinin was previously reported (Adair, W. S., C. J. Hwang, and U. W. Goodenough, 1983, Cell, 33:183-193). Here we characterize the purified minus molecule. We show it to be a high molecular weight, hydroxyproline-rich glycoprotein that migrates in the 3% stacking region of an SDS-polyacrylamide gel and is absent from two nonagglutinating minus mutants. Plus and minus agglutinins are remarkably similar, although nonidentical, in amino acid composition, molecular morphology, and reactivity in vivo and in vitro with monoclonal antibodies raised against the plus agglutinin. Moreover, the adhesiveness of both plus and minus agglutinins, when coupled to agarose beads, is abolished by thermolysin, trypsin, periodate, alkaline borohydride, reducing agents, or heat, but unaffected by exo- or endoglycosidases. The minus agglutinin, however, migrates just ahead of the plus molecule on SDS PAGE, is excluded from an anion-exchange (Mono Q) column, elutes earlier during hydrophobic interaction (Bio-gel TSK Phenyl 5PW) chromatography, and is sensitive to chymotrypsin digestion (unlike the plus agglutinin); therefore, it differs from the plus agglutinin in apparent molecular weight, net charge, relative hydrophobicity and proteolytic susceptibility. Nevertheless, our results generally demonstrate a high degree of homology between these complementary cell-cell recognition/adhesion molecules, which suggests that they are specified by genes that have a common evolutionary origin.
 ...
PMID:Characterization of the purified Chlamydomonas minus agglutinin. 241 36

The positions of the inter- and intra-chain disulfide bonds of human plasma alpha 2 HS-glycoprotein were determined. alpha 2 HS-glycoprotein was digested with acid proteinase and then with thermolysin. The disulfide bonds containing peptides were separated by reversed-phase HPLC and detected by SBD-F (7-fluorobenzo-2-oxa-1,3-diasole-4-sulfonic acid ammonium salt) method. One inter-disulfide bond containing peptide and five intra-disulfide bond containing peptides (A-chain) were purified and identified as Cys-18 (B-chain)--Cys-14 (A-chain), Cys-71--Cys-82, Cys-96--Cys-114, Cys-128--Cys-131, Cys-190--Cys-201 and Cys-212--Cys-229, respectively. The location of the intra-disulfide bonds revealed that the A-chain of alpha 2 HS-glycoprotein is composed of three domains. Two domains were shown to possess intramolecular homology judging from the total chain length of the domains, size of the loops formed by the S--S bonds, the location of two disulfide loops near the C-terminal end of domains A and B, the distance between two S--S bonds of each domain, the amino acid sequence homology between these two domains (22.6%), number of amino acid residues between the second S--S loops and the end of domains A and B, and the positions of the ordered structures.
 ...
PMID:The position of the disulfide bonds in human plasma alpha 2 HS-glycoprotein and the repeating double disulfide bonds in the domain structure. 264 41


1 2 3 4 5 Next >>