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

The hydrolysis of a series of depsipeptides demonstrates that the zinc neutral endopeptidases of bacteria are active esterases. Esters such as BzGly-OPhe-Ala, BzGly-OLeu-Ala, and FA-Gly-OLeu-NH2 are hydrolyzed at rates three- to eightfold slower than are their exact peptide analogues, when hydrolyzed by thermolysin, Bacillus subtilis neutral protease and the neutral protease from Aeromonas proteolytica. Ester hydrolysis by zinc neutral proteases follows the characteristic preference for hydrophobic amino acids adjacent to the site of cleavage, discerned from the hydrolysis of peptide substrates. Removal of zinc from thermolysin abolishes the esterase activity of the native enzyme. Among the metals examined, only Co2+ and Zn2+ restore esterase activity to any significant extent, Co2+ restoring 50% and Zn2+ 100% of the native thermolysin activity. The hydrolysis of esters and peptides by thermolysin does not differ with respect to either the binding or catalytic steps. Substrate specificity, pH-rate profiles, inhibitor, and deuterium isotope effects are identical for both types of substrates.
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PMID:Esterase activity of zinc neutral proteases. 0 76

A carboxypeptidase was purified to electrophoretic homogeneity from the thermoacidophilic archaebacterium Sulfolobus solfataricus. Molecular masses assessed by SDS/PAGE and gel filtration were 42 kDa and 170 kDa, respectively, which points to a tetrameric structure for the molecule. An isoelectric point of 5.9 was also determined. The enzyme was proven to be a metalloprotease, as shown by the inhibitory effects exerted by EDTA and o-phenanthroline; furthermore, dialysis against EDTA led to a complete loss of activity, which could be restored by addition of Zn2+ in the micromolar range, and, to a lesser extent, by Co2+. The enzyme was endowed with a broad substrate specificity, as shown by its ability to release basic, acidic and aromatic amino acids from the respective benzoylglycylated and benzyloxycarbonylated amino acids. An esterase activity of the carboxypeptidase was also demonstrated on different esterified amino acids and dipeptides blocked at the N-terminus. The enzyme displayed broad pH optima ranging over 5.5-7.0, or 5.5-9.0, when using an acidic or a basic benzyloxycarbonylated amino acid, respectively. With regard to thermostability, it was proven to be completely stable on incubation for 15 min at 85 degrees C. Furthermore, thanks to its relatively low activation energy, i.e. 31.0 kJ/mol, it was still significantly active at room temperature. At 40 degrees C, the enzyme could withstand 0.1% SDS and different organic solvents: particularly ethanol up to 99%. Amino acid and N-terminal sequence analyses did not evidence any similarity to carboxypeptidases A nor thermolysin. A weak similarity was only found with bovine carboxypeptidase B.
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PMID:Purification and characterization of a thermostable carboxypeptidase from the extreme thermophilic archaebacterium Sulfolobus solfataricus. 159 79

Two glycoproteins having trypsin-protein esterase activity were purified to apparent homogeneity from murine plasma. One was alpha-macroglobulin, a homologue of human alpha-2-macroglobulin, while the other, tentatively named murinoglobulin, did not correspond to any of the known plasma protease inhibitors that have been well characterized in men or other mammals. Murinoglobulin contained about 7.6% carbohydrate and was composed of a single-polypeptide chain of Mr = 180,000 as judged by the equilibrium sedimentation analysis and sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing conditions. Murinoglobulin did not cross-react immunologically with mouse alpha-macroglobulin nor with human alpha-2-macroglobulin. Protease-inhibiting properties of murinoglobulin were compared with those of mouse alpha-macroglobulin and human alpha-2-macroglobulin. All the three proteins inhibited trypsin, papain, and thermolysin, although they differed considerably in both the degree of inhibition and the binding stoichiometry of protease-inhibitor complexes. The two macroglobulins inhibited pepsin at pH 5.5, whereas murinoglobulin was inactivated at this pH. Murinoglobulin was more sensitive to methylamine than the two macroglobulins. No protein corresponding to murinoglobulin was detected in human plasma.
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PMID:Murinoglobulin, a novel protease inhibitor from murine plasma. Isolation, characterization, and comparison with murine alpha-macroglobulin and human alpha-2-macroglobulin. 257 55

Ovalbumin is partially homologous in sequence with the proteinase inhibitors alpha 1-proteinase inhibitor and anti-thrombin III. The region of sequence in ovalbumin which corresponds to the reactive sites of these proteinase inhibitors is susceptible to attack by subtilisin, elastase, thermolysin, bromelain, and Bacillus cereus protease. The esterase activity of elastase is not inhibited by ovalbumin, but ovalbumin is efficiently cleaved by elastase. In contrast with these proteases, trypsin does not cleave ovalbumin.
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PMID:Ovalbumin is an elastase substrate. 656 26

Several proteases were studied as potential catalysts for the enzymatic synthesis of oligopeptides containing the unnatural amino acid allylglycine, the overall objective being the synthesis of a reactive tetrapeptide that could be chemically polymerized into a potentially biocompatible or biodegradable material. Commercially available enzymes were screened for esterase activity toward the methyl ester of the amino acid allylglycine (DL-AgOMe) to identify potential catalysts for dipeptide synthesis. Proteases from Aspergillus oryzae and Aspergillus sojae, pronase E and protease Nagarse synthesized the protected dipeptide Cbz-allylglycine-phenylalaninamide (Cbz-L-Ag-L-PheNH2) from Cbz-DL-AgOMe and L-PheNH2. However, the same enzymes were not able to catalyze the synthesis of Cbz-phenylalanine-allylglycine ethyl ester (Cbz-L-Phe-L-AgOEt). Thus, although these enzymes could use allylglycine as the acyl donor they could not employ it as the acyl acceptor in peptide synthesis. In contrast, chymotrypsin was able to use allylglycine ethyl ester (DL-AgOEt) as the acyl acceptor in the synthesis of Cbz-L-Phe-L-AgOEt, but was not able to synthesize Cbz-L-Ag-L-PheNH2. The two dipeptides, Cbz-allylglycine-phenylalanine and phenylalanine-allylglycine ethyl ester, served as substrates for the thermolysin-catalyzed synthesis of the tetrapeptide Cbz-L-Ag-L-Phe-L-Phe-L-AgOEt.
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PMID:Enzymatic synthesis of peptides containing unnatural amino acids. 854 Oct 21

We report here the expression of recombinant human prokallikrein and kallikrein in engineered Chinese hamster ovary cells transfected with a human genomic gene encoding preprokallikrein. At high expression levels, recombinant prokallikrein, an inactive proenzyme form, is predominantly secreted into the culture medium. Upon chromatographic separations, the inactive prokallikrein as well as the mature kallikrein after thermolysin activation of the proenzyme can be prepared to apparent purity. Both prokallikrein and kallikrein can be further separated into two distinct high- and low-molecular-weight isoforms. Kallikrein preparations are fully active in standard kallikrein activity assays such as esterase activity and kinin release from kininogen. Both kallikrein and prokallikrein display multiple molecular forms with differences in both molecular sizes and charges. The structural differences in high- and low-molecular-weight kallikreins or prokallikreins were found to be due to glycosylation, with the high-molecular-weight species glycosylated at three Asn-linked sites and the low-molecular-weight species at two of the three Asn-linked sites. The multiply charged kallikrein isoforms are derived from different numbers of sialic acids attached at the detected Asn-linked carbohydrates. In comparison with kallikrein, prokallikrein appears to show a significant decrease in the magnitude of near uv-circular dichroism bands, suggesting a change in local conformation. This conformational change correlates with the loss of activity in proenzyme due to the presence of propeptide.
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PMID:Purification and characterization of human tissue prokallikrein and kallikrein isoforms expressed in Chinese hamster ovary cells. 881 67

A novel thermostable neutral proteinase, called NPS, was purified to electrophoretic homogeneity from the culture broth of Saccharomonospora canescens sp. novus, strain 5. The molecular mass was determined by SDS-polyacrylamide gel electrophoresis to be 35,000 Da. The enzyme exhibits a sharp pH optimum of proteolytic activity at pH 6.7. NPS was completely inactivated with inhibitors, typical for metalloendopeptidases, EDTA and 1,10-phenantroline, whereas the serine proteinase inhibitor PMSF had no effect. Atomic absorption measurements showed that the proteinase binds a single zinc and four calcium ions. The enzyme thermostability was characterized in the absence and presence of added calcium. Melting temperature, Tm = 77 degrees C and an activation energy, Ea, for the thermal deactivation of the excited protein fluorophores of 72.13 kJ mol-1 were calculated in the presence of 100 mM CaCl2. The Ea-value is considerably higher than those obtained for a number of proteinases from microorganisms and was explained by the thermostable structure of the enzyme. Effective radiationless energy transfer from phenol groups to indole rings was observed. 68% of the light absorbed by tyrosyl residues is transferred to tryptophyl side chains. No homology was found after comparison of the NPS N-terminal sequence, including the first 26 residues, with those of other neutral proteinases from microorganisms. In contrast to the well-known bacterial neutral proteinase thermolysin and related enzymes from microorganisms, NPS possesses arylamidase and esterase activities. Further crystallographic studies will reveal the structural reasons for this specificity. Epoxy and epithio pyranosides are inhibitors of the proteinase arylamidase activity.
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PMID:A novel thermostable neutral proteinase from Saccharomonospora canescens. 954 Jul 92

For (S)-thiirancarboxylic acid a second-order rate constant of k2nd = 222 M(-1) min(-1) for the irreversible inhibition of papain was determined. The ethyl and methyl ester do not inhibit the enzyme time-dependently. An improved synthesis of enantiomerically pure thiirancarboxylic acid is described. It is shown that thiirancarboxylates can be substrates for serine proteases (alpha-chymotrypsin) and esterases (pig liver esterase) and even for metallo proteases (thermolysin).
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PMID:(S)-Thiirancarboxylic acid as a reactive building block for a new class of cysteine protease inhibitors. 1112 43