Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.21.1 (chymotrypsin)
 10,938 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A basic protein (pI 10.2), named basic protein I, was purified to homogeneity from the venom of Trimeresurus flavoviridis (Habu snake) after four chromatographic steps. The amino acid sequence of this protein was determined by sequencing the S-pyridylethylated derivative of the protein and its peptides produced by chemical (cyanogen bromide and formic acid) and enzymatic (chymotrypsin, Achromobacter protease I, and Staphylococcus aureus V8 protease) cleavages. The protein consisted of 122 amino acid residues and was similar in sequence to phospholipases A2 from the venoms of crotalid and viperid snakes. A most striking feature of this protein is that aspartic acid at the 49th position common in phospholipases A2 is replaced by lysine. When the protein acted on 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphorylcholine, oleic acid was preferentially released, indicating that the protein has phospholipase A2 activity. Its molar activity toward 1,2-dilauroyl-sn-glycero-3-phosphorylcholine, however, was 1.5% that of T. flavoviridis phospholipase A2 isolated previously. The fact that both affinity to Ca2+ and reactivity to p-bromophenacyl bromide of basic protein I are approximately one order of magnitude lower than those of T. flavoviridis phospholipase A2 might explain the low activity of basic protein I.
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PMID:Purification and amino acid sequence of basic protein I, a lysine-49-phospholipase A2 with low activity, from the venom of Trimeresurus flavoviridis (Habu snake). 233 Jun 4

A peptide having enzyme-like catalytic activity has been designed and synthesized. Computer modeling was used to design a bundle of four short parallel amphipathic helical peptides bearing the serine protease catalytic site residues serine, histidine, and aspartic acid at the amino end of the bundle in the same spatial arrangement as in chymotrypsin (ChTr). The necessary "oxyanion hole" and substrate binding pocket for acetyltyrosine ethyl ester, a classical ChTr substrate, were included in the design. The four chains were linked covalently at their carboxyl ends. The peptide has affinity for ChTr ester substrates similar to that of ChTr and hydrolyzes them at rates approximately 0.01 that of ChTr; total turnovers greater than 100 have been observed. The peptide is inhibited by ChTr specific inhibitors and is inactive toward benzoyl arginine ethyl ester, a trypsin substrate. The peptide is inactivated by heating above 60 degrees C, but recovers full catalytic activity upon cooling and lyophilization from acetic acid.
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PMID:Design and synthesis of a peptide having chymotrypsin-like esterase activity. 236 48

The dimeric association process of alpha-chymotrypsin has been studied with the aid of a stopped-flow spectrophotometer at various temperatures and pH values. From the temperature dependences of the forward reaction rate constant (kf) and the equilibrium dimerization constant (KD), the reaction system observed here is concluded to be entropy-driven. The increase in entropy can be attributed to the release of water molecules from both the active site and the surface part of the protein molecule during the course of dimerization. From the pH dependences of the reaction rate constants and the equilibrium constant, the reaction is concluded to depend strongly on the dissociations of the site between the carboxyl group of the aspartic acid and imidazolyl group of the histidine residues (in the higher pH region), and the site between the imidazolyl group of the histidine and the carboxyl group of the tyrosine residue (in the lower pH region), respectively.
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PMID:Kinetic study of the effects of solvation on the dimerization process of alpha-chymotrypsin. 272 89

The effects of poly(L-aspartic acid) (PLAA) on microtubule assembly and microtubule-associated protein (MAP) 2-actin interaction were examined in vitro. PLAA inhibited assembly of rat brain microtubules and induced rapid disassembly of already formed microtubules. Inhibition was stronger by PLAA with a high molecular weight than that by low molecular weight. The ratios of 47 kDa PLAA to microtubule proteins causing 50% inhibition of the assembly and disassembly were 0.015 and 0.04 (w/w), respectively. Both MAP 1 and MAP 2 were bound to a PLAA-Sepharose 4B affinity column, while tubulin was not retained by the column. PLAA caused selective dissociation of MAP 1 and MAP 2 from microtubules polymerized by taxol. It is therefore concluded that PLAA interacts specifically with MAPs. PLAA also inhibited the MAP 2 induced cross-linking of actin filaments, showing an almost complete inhibition at a PLAA to MAP 2 ratio of 1:5,000 (w/w). Binding experiments of PLAA with digested MAP 2 by chymotrypsin using affinity chromatography and sedimentation experiments showed that PLAA was preferentially bound to a 35 kDa fragment which includes the microtubule- and actin-binding domain of the MAP 2 molecule. These results suggest that PLAA suppressed the functions of MAP 2 through a domain which is located in the 35 kDa fragment.
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PMID:Inhibitory effects of poly(L-aspartic acid) on the assembly of brain microtubules and the interaction of microtubule-associated protein 2 with F-actin in vitro. 277 57

The steroidal 3 beta-oxirane (3S)-spiro[5 alpha-androstane-3,2'-oxiran]-17 beta-ol (1 beta) is an active site directed irreversible inhibitor of the 3-oxo-delta 5-steroid isomerase from Pseudomonas testosteroni. Two steroid-bound peptides (TPS1 and TPS2) were isolated by high-performance liquid chromatography (HPLC) from the trypsin digest of enzyme inactivated with 1 beta. The modified tryptic peptides (residues 14-45 of the enzyme) were further digested with chymotrypsin, each giving rise to a single steroid-containing product (CPS1 and CPS2, respectively) derived from residues 31 to 45 of the enzyme. The modified chymotryptic peptides were isolated by HPLC, and the peptide-steroid ester linkage was reduced with sodium hydroxyborohydride. Amino acid analysis of the reduced peptides gave ca. 0.5 residue of homoserine and one less residue of aspartic acid than the corresponding unreduced peptides. Sequence analysis of both reduced chymotryptic peptides revealed that homoserine was located at position 8 in the peptide sequence, corresponding to residue 38 of the enzyme. The finding that the steroidal 3 beta-oxirane, like the 17 beta-oxiranes, inactivates the isomerase via esterification of aspartic acid-38 is strong evidence that this enzyme binds steroids in at least two orientations.
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PMID:Affinity alkylation of 3-oxo-delta 5-steroid isomerase by steroidal 3 beta-oxiranes: identification of the modified amino acid by reduction with hydroxyborohydride. 362 Apr 46

This report describes the purification of a novel proteinase inhibitor from bovine serum. This protein was purified to apparent homogeneity employing affinity binding to sulfated dextran and precipitation by ammonium sulfate, followed by sequential chromatography on DEAE-cellulose, heparin-Sepharose and Sephacryl S-200. Quantitative enzyme-linked immunosorbent assays revealed that the concentration of this inhibitor is approximately 3 microM in bovine serum. The inhibitor is a single polypeptide chain with an estimated Mr of 83,000 as determined by SDS-polyacrylamide gel electrophoresis. An aspartic acid was found at the amino terminus of the protein; N-terminal amino acid sequence data indicated that there was no significant homology with other reported amino acid sequences. This bovine inhibitor covalently complexed the human proteinases C1-r, C1-s, factor XIIa and plasma kallikrein, which are also complexed and inactivated by human C1-inhibitor. In addition, the bovine inhibitor complexed and inactivated bovine chymotrypsin, a feature which functionally distinguishes it from human C1-inhibitor. Although the bovine inhibitor appears functionally very similar to C1-inhibitor, we found no evidence for structural homology with the human counterpart.
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PMID:Purification of a proteinase inhibitor from bovine serum with C1-inhibitor activity. 381 12

Double-headed protease inhibitors I, IIa, and IIc (AB I, AB IIa, and AB IIc) have been purified from azuki beans "Takara" (Vigna angularis) by conventional chromatographic methods and their amino acid sequences have been determined. AB I, AB IIa, and AB IIc had molecular weights of 9,166, 8,661, and 8,756 daltons, consisting of 82, 78, 79 amino acid residues, respectively. The molecular weights of these inhibitors, determined by gel filtration at pH 8.0, were 18,000 for AB I and 17,000 for both AB IIa and AB IIc, indicating that the inhibitors are dimers. The inhibitors had isoelectric points of 4.7 (AB I), 6.8 (AB IIa), and 6.2 (AB IIc). AB I stoichiometrically inhibited both trypsin and chymotrypsin at a molar ratio of 1 : 1. On the other hand, AB IIa and AB IIc both inhibited trypsin at a molar ratio of about 1 : 2 and also inhibited chymotrypsin, though only weakly. Sequence comparison with other double-headed inhibitors indicated the reactive sites of AB IIa and AB IIc for trypsin to be Lys26-Ser27 and Arg53-Ser54, and those of AB I for trypsin and chymotrypsin to be Lys26-Ser27 and Tyr53-Ser54, respectively. The differences between AB IIa and AB IIc were that AB IIa lacked the C-terminal aspartic acid residue, and that Glu10 and Arg60 in AB IIa were replaced by Gln10 and His60 in AB IIc. A comparison between AB IIa and AB I revealed 25 variant amino acids among the 78 residues of AB IIa; further, Ab IIa lacked 4 amino acid residues in the C-terminal region of AB I.
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PMID:Azuki bean (Vigna angularis) protease inhibitors: isolation and amino acid sequences. 388 75

We previously have isolated cDNA clones for several mRNAs that increase in abundance during the differentiation of 3T3 adipocytes but whose physiological role is unknown. We show here that a mRNA that is complementary to one of these clones and encodes a protein of 28 kDa is expressed abundantly in mouse fat pads but not in several other mouse tissues. Sequence analysis of the corresponding cDNA clone indicated that the encoded protein shows 30% overall amino acid homology to several serine proteases including trypsin, chymotrypsin, and elastase. Homology is much higher (64%) between the 28-kDa protein and regions that are strongly conserved among the members of the serine protease family. The derived protein also has key features characteristic of active serine proteases, including the histidine, aspartic acid, and serine residues, which comprise the charge relay system, and a potential cleavage site for activation of the zymogen. Primer extension analysis performed to obtain the sequence of the 5' end of mRNA that encodes the 28-kDa protein indicates that two forms of this mRNA exist and probably arise through alternative splicing. The two mRNAs encode signal sequences that differ by the deletion of one amino acid near the predicted cleavage site of the signal peptide. These results demonstrate that adipocyte differentiation is accompanied by the expression of mRNA encoding a serine protease homologue that can be synthesized with two different signal peptides.
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PMID:A developmentally regulated mRNA from 3T3 adipocytes encodes a novel serine protease homologue. 390 Oct 3

Proton NMR spectra of serine proteases in 1H2O solutions typically show a single resonance at very low magnetic field--i.e., 14-18 ppm from dimethylsilylapentanesulfonate. This resonance has been assigned to the proton hydrogen bonded between aspartic acid-102 and histidine-57 (chymotrypsin numbering system) of the "charge-relay system" or catalytic triad of serine proteases [Robillard, G. & Shulman, R. G. (1972) J. Mol. Biol. 71, 507-511]. Since then, there have been a number of reports that have cast doubt on its correctness. In the present work we have tested this assignment using alpha-lytic protease (EC 3.4.21.12, Myxobacter alpha-lytic proteinase), a bacterial serine protease homologous to elastase, which is specifically labeled with nitrogen-15 at N delta 1 of its single histidine residue. The low-field region of the proton spectra of this labeled enzyme shows a single resonance having the properties reported [Robillard, G. & Shulman, R. G. (1974) J. Mol. Biol. 86, 519-540], which, in addition, exhibits spin-spin splitting to the nitrogen-15 label. The observation of this 15N delta 1-H coupling makes the assignment of this resonance to the charge-relay proton unequivocal.
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PMID:Confirmation of the assignment of the low-field proton resonance of serine proteases by using specifically nitrogen-15 labeled enzyme. 393 65

Two forms of hen ovalbumin that exist as genetic variants were compared by physical and immunochemical techniques. The two ovalbumins could be distinguished by electrophoretic mobility and by antisera that had been pretreated with heterologous antigen. Peptide ;maps' of chymotrypsin digests of the two ovalbumins revealed that three of the peptides were different. These were isolated and analysed. One form of ovalbumin (type B) contains the sequence: -Ser-Ser-Ala-Asp-Leu-Ser-Gly-Ile-Ala-Glu-Ser(Ser,Leu)- whereas the other form (type A) contains an asparagine residue in place of the aspartic acid residue. The -Asn-Leu-Ser- sequence of type A is not glycosylated. Chymotrypsin readily cleaved the leucine-serine bond in the asparagine-containing peptide, but not in the aspartic acid-containing variant.
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PMID:Replacement of asparagine by aspartic acid in hen ovalbumin and a difference in immunochemical reactivity. 507 26


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