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

An acid proteinase from Aspergillus oryzae was isolated from a commercial powder by successive (NH4)2SO4 fractionation, acetone precipitation, and ion-exchange chromatography on phosphate- and DEAE-cellulose columns. The purified enzyme was found to be homogeneous by ultracentrifuge-sedimentation analysis (S20, W equal 3.63S), but electrofocusing in polyacrylamide gels and electrophoresis at pH 3.2 revealed that it consists of two very closely migrating bands. No difference in the amino acid composition and enzymic activities of the two partially separated bands could be detected, and it was concluded that the acid proteinase exists in two molecular forms. The enzyme activates bovine trypsinogen and chymotrypsinogen at pH 3.5 (the kappacat. and Km values at 35degrees C are 11.3S- minus 1, 0.10mM and 1.14S- minus 1, 0.18mM respectively). It hydrolyses the Phe-Phe bond of the synthetic pepsin substrates Z-His-Phe-Phe-OEt (kappacat. equal 1.65S- minus 1, Km equal 0.640mM at pH 3.5, 30degrees C) and Z-Ala-Ala-Phe-Phe-OPy4Pr (kappacat. equal 0.37S- minus 1, Km equal 0.037 mM at pH2.9, 39degrees C), where Z represents benzyloxycarbonyl and OPy4Pr represents 3-(4-pyridyl)-propyl 1-ester. Activation of bovine chymotrypsinogen results from the cleavage of the Arg(15)-Ile(16) bond in the zymogen. No other cleavages were observed. The use of A. oryzae proteinase provides a simple tool for the production of pi-chymotrypsin in good yield and purity.
 ...
PMID:Aspergillus oryzae acid proteinase. Purification and properties, and formation of pi-chymotrypsin. 23 2

The steady-state kinetics of plasmin (EC 3.4.21.7) catalysed reactions with some alpha-N-benzoyl-L-arginine compounds is investigated in the pH range 5.8--9.0. The results are interpreted in terms of a three-step mechanism, which involves enzyme-substrate complex formation, followed by acylation and deacylation of the enzyme. Alpha-N-Benzoyl-L-arginine methyl ester and ethyl ester show the same pH behaviour. The kinetic parameter kc/Km is influenced by two groups with pK values of 6.5 and 8.4, respectively. kc is affected only by the group with pK equal to 6.5 and Km only by the group with pK equal to 8.4. It is suggested that the group with pK equal to 6.5 is the 1-chloro-3-tosyl-amido-7-amino-2-heptanone-sensitive histidine residue in the active site and that the group with pK equal to 8.4 is perhaps the alpha-amino group of the N-terminus in analogy to trypsin and chymotrypsin. alpha-N-Benzoyl-L-arginine amide is not hydrolysed by plasmin, but proves to be a competitive inhibitor, Ki = 12.8 +/- 1.8 mM, pH = 7.8. Also the product alpha-N-benzoyl-L-arginine is a competitive inhibitor, Ki = 26 +/- 3.1 mM, pH = 7.8. Estimates of individual rate constants are compared with similar trypsin data.
 ...
PMID:pH effects in plasmin-catalysed hydrolysis of alpha-N-benzoyl-L-arginine compounds. 23 53

The use of a linear free-energy relationship shows that both histidine residues of alpha-chymotrypsin and chymotrypsinogen are super-reactive toward 1-fluoro-2,4-dinitrobenzene. The binding of indole to the specificity site of alpha-chymotrypsin causes both histidine residues to become less reactive. On the basis of these results and those from X-ray-crystallographic studies, the following conclusions are made. (1) The super-reactivity of the catalytic-site histidine-57 is due to charge transfer from aspartic acid-102 by means of hydrogen bonding. (2) The aspartic acid-102-histidine-57-serine-195 'charge-relay' system is not complete in the zymogen or native enzyme and only on binding of a suitable substrate or ligand to the specificity site of the enzyme is the charge transfer to serine-195 completed. (3) The lack of substantial enzymic activity in the zymogen is due to the absence of a completed specificity site, and therefore it cannot bind suitable substrates or ligands to induce completion of the charge-relay system.
 ...
PMID:Properties of the histidine residues of indole-chymotrypsin. Implications for the activation process and catalytic mechanism. 24 27

This paper presents the experimental details which led to the elucidation of the complete primary structure of S16, a protein which belongs to the small subunit of E. coli ribosomes. Protein S16 was digested with trypsin, alpha-chymotrypsin, and the staphylococcal protease. The resulting peptides were purified on paper and their amino acid composition and sequence were determined. Automatic Edman degradation with a modified sequenator on the complete protein yielded information from the 56N-terminal residues. The combination of all these results led to the following complete amino acid sequence: Met-Val-Thr-Ile-Arg-Leu-Ala-Arg-His-Gly-Ala-Lys-Lys-Arg-Pro-Phe-Tyr-Gln-Val-Val-Val-Ala-Asp-Ser-Arg--Asn-Ala-Arg-Asn-Gly-Arg-Phe-Ile-Glu-Arg-Val-Gly-Phe-Phe-Asn-Pro-Ile-Ala-Ser-Glu-Lys-Glu-Glu-Gly-Thr-Arg-Leu-Asp-Leu-Asp-Arg-Ile-Ala-His-Trp-Val-Gly-Gln-Gly-Ala-Thr-Ile-Ser-Asp-Arg-Val-Ala-Ala-Leu-Ile-Lys-Glu-Val-Asn-Lys-Ala-Ala. The molecular weight derived from the sequence amounts to 9 162.
 ...
PMID:The complete amino acid sequence of protein S16 from Escherichia coli. 33 10

Phenylalanine chloromethyl ketone covalently attached to porous glass beads was synthesized to serve as a solid-phase active site directed inhibitor of chymotrypsin-like proteolytic enzymes. The solid-phase reagent inhibited 20 nmol of bovine chymotrypsin per gram of glass and covalently bound 30 nmol of protein per gram of glass. Sepharose-bound lysine chloromethyl ketones were synthesized to serve as inhibitors of trypsin-like enzymes. Sepharose-MethionylLysyl chloromethyl ketone inactivated and bound about 6.8 nmol of enzyme per ml of settled gel. In a preliminary experiment, a cyanogen bromide cleavage of the methionine residues showed that it should be possible to release all peptides but the peptide containing the active-site histidine. The immobilized trypsin was also reduced, carboxymethylated and digested with chymotrypsin. The potential of the solid-phase approach is in the isolation of a specific serine proteinase and in the sequence determination of residues surrounding the active-site histidine.
 ...
PMID:Solid-phase active site inhibitors of proteinases. 42 26

The trypsin inhibitors from winged bean seed were isolated by affinity chromatography on trypsin-Sepharose 4B and the components fractionated by chromatography on SP-Sephadex C-25 and Sephadex G-100. The major components, inhibitors 2 and 3 were found to be homogeneous proteins with molecular weights of about 20,000. The inhibitors stoichiometrically inhibited bovine trypsin in the molar ratio of 1 : 1 whereas the inhibition of bovine alpha-chymotrypsin was weak and non-stoichiometric. Amino acid analysis indicated that both the inhibitors contain four cysteine residues and are rich in aspartic acid, glutamic acid, glycine, valine and leucine; however, inhibitor 3 lacks histidine and methionine while inhibitor 2 contains one histidine and three methionines. A minor trypsin inhibitor fraction was also isolated which contained at least three proteins with a molecular weight of about 10,000 and a high content of half-cystine.
 ...
PMID:Isolation and characterization of the trypsin inhibitors from winged bean seed (Psophocarpus tetragonolobus (L) Dc.). 45 47

The amino acid sequence of the beta-chain of the principal haemoglobin from the shark H. portusjacksoni has been determined. The chain has 141 residues, the same as that of mammalian alpha-chains and less than the 146 residues of mammalian beta-chains or the 148 residues of the alpha-chain from the tetrameric shark haemoglobin. The sequence was deduced from the sequences of peptides obtained by digestion of the globin or its cyanogen bromide fragments with trypsin, chymotrypsin, pepsin and papain. The difference in length of the beta-chain is most readily accounted for by the absence of the D helix. This small helical section is normally present in myoglobins and beta-globins but absent in alpha-chains. The deduction that it is absent from shark beta-chain is based on consideration of homology. The beta-chain shows the insertion of histidine beta2 and the deletions corresponding to residues A17 and AB1 relative to alpha-and myoglobin chains. The reactive thiol group in shark haemoglobin was shown by radioactive labelling to be residue 51 in the beta-chain, immediately preceding the E helix. The amino acid sequence of shark beta-chain shows 92 differences from human beta-chain, significantly more differences than shown by chicken or frog beta-chains, in line with its earlier time of divergence. If the tertiary structure of the shark beta-chain is the same as that of the horse then there are two changes in the alpha1beta2 contact site in oxyhaemoglobin and an additional one in deoxyhaemoglobin. When both alpha- and beta-chain contacts are considered there is a total of nine changes in residues involved in the alpha1beta2 contacts. There is no Bohr effect in shark haemoglobin, and of the residues normally involved in this effect the C-terminal histidine residue of the beta-chain is present, but the aspartyl (FG1) residue to which it is salt-linked is not, being replaced by a glutamyl residue.
 ...
PMID:Haemoglobins of the shark, Heterodontus portusjacksoni. III. Amino acid sequence of the beta-chain. 61 4

The effect of methylation of histidine-57 of alpha-chymotrypsin with Streptomyces subtilisin inhibitor was examined. Methylchymotrypsin was isolated by affinity chromatography on inhibitor-Sepharose, and the interaction of this inactive enzyme with inhibitor was quantitatively analyzed by two different methods: the spectrophotometric titration of difference spectrum resulted in the complex formation and the application of competitive enzyme assay by using substrates of large Km values. The former method gave values of 8.6 . 10(-6) M as dissociation constant (Kd) of methylchymotrypsin . inhibitor complex and 0.91 as the number of binding sites (n) per inhibitor monomer, both of which were almost equivalent to those for native enzyme . inhibitor complex. By the latter novel method, values of 7.9 . 10(-6) M and 1.08 were obtained for Kd and n, respectively, for interaction of inhibitor with alpha-chymotrypsin, and 8 . 10(-6) M as Kd for methylchymotrypsin . inhibitor complex. These results indicate that methylation of histidine-57 of active site in alpha-chymotrypsin molecule does not affect essentially the binding ability to inhibitor and the modified enzyme binds stoichiometrically to inhibitor, as the native enzyme does, with a molar ratio of 1:1 per inhibitor monomer.
 ...
PMID:Interaction of methylchymotrypsin with Streptomyces subtilisin inhibitor. 62 38

"Group-specific" protease (GSP) is a serine protease, obtained from rat small intestine, which preferentially inactivates the apo forms of certain pyridoxal phosphate requiring enzymes. The enzyme contains 224 amino acid residues in a single polypeptide chain and three disulfide bonds. In the present work the covalent structure has been determined and its homologous relationship to those of chymotrypsin, trypsin, and elastase has been established (approximately 33% identity with each). The residues forming the "charge-relay" system of the active site of chymotrypsin (His-57, Asp-102, and Ser-195) are found in corresponding regions in GSP, whereas an alanyl residue at position 176 of GSP corresponds to a residue which participates in the primary substrate binding site in serine proteases (Asp-177 in trypsin; Ser-189 in chymotrypsin). Three disulfide bonds in GSP occur in similar positions in chymotrypsin, trypsin, and elastase. However, GSP lacks a disulfide bond which is present in all known serine proteases (linking Cys-191 to Cys-220 in chymotrypsin). In view of the close proximity of this bond to both the primary and the antiparallel binding sites of various serine proteases, it is likely that its absence in GSP is related to the substrate specificity of this enzyme. It is concluded that GSP diverged from a common ancestor preceding chymotrypsin but following trypsin.
 ...
PMID:Covalent structure of a group-specific protease from rat small intestine. Appendix: crystallographic data for a group specific protease from rat intestine. 62 33

The one-electron reduction of chymotrypsin, trypsin, and their zymogens have been studied by pulse radiolysis. The optical spectra of the transient products from the two active enzymes display a pH-dependent band at 360 nm, associated with the histidine-electron adduct. The yield of the histidyl radical as a function of pH is consistent with a pK(a) less than 4.5, which suggests that the radical is located at the enzyme active site. The histidines of the proenzymes chymotrypsinogen and trypsinogen are unreactive towards the hydrated electron. We conclude that formation of the histidine-electron adduct at the serine protease active site is sensitive to the physical alterations which accompany protease activation.
 ...
PMID:Application of pulse radiolysis to the study of proteins: chymotrypsin and trypsin. 70 36


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>