Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.4 (trypsin)
42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A new ATPase electrophoretically and immunologically distinct from the dynein ATPase studied previously has been solublized and purified from sea urchin sperm flagella. This ATPase has properties similar to those of dynein ATPase. Therefore, we propose that the two ATPases be considered as dynein isoenzymes, with previously studied dynein being known as dynein 1, and the newly discovered ATPase as dynein 2. Some physicochemical and enzymatic properties of dynein 2 have been determined. The molecular weight calculated from the sedimentation coefficient (12.3 "/- 1 S) and Stokes radius (12.8 "/- 0.4 nm) is 690,000 +/- 70,000. The molecular weight of the high molecular weight subunit of dynein 2 has been determined to be 325,000 +/- 40,000 by Na dodecyl-SO4-polyacrylamide gel electrophoresis. The enzymatic properties of dynein 1 and dynein 2 are similar in substrate specificity, pH optimum, and Mg2+ requirement for ATPase activity, but they differ in their Michaelis constant and in their dependence of ATPase activity upon salt concentration. Digestion of dynein 2 with trypsin yields an ATPase-containing protein fragment, similar to Fragment A obtained from dynein 1. An antiserum prepared against Fragment A from dynein 1 did not precipitate dynein 2 or inhibit its ATPase activity.
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PMID:Dynein 2. A new adenosine triphosphatase from sea urchin sperm flagella. 13 96

A method is described for the simultaneous purification of milligram quantities of complement components C2 and Factor B. Both products are homogeneous by the criteria of polyacrylamide-gel electrophoresis and N-terminal sequence analysis. Component C2 is cleaved by serine proteinase C1s at an X-Lys bond to give fragment C2a (approx. mol.wt. 74000) and fragment C2b (approx. mol.wt. 34000). The two fragments can be separated by gel filtration without the need for reducing or denaturing agents. Fragment C2b represents the N-terminal end of the molecule. Similar results were seen on cleavage of Factor B by Factor D in the presence of component C3. Again two non-covalently linked fragments are formed. The smaller, fragment Ba (approx. mol.wt. 36,000),) has threonine as the N-terminal residue, as does Factor B; the larger, fragment Bb (approx. mol. wt. 58000), has lysine as the N-terminal residue. A similar cleavage pattern is obtained on limited proteolysis of Factor B by trypsin, suggesting an Arg-Lys-or Lys-Lys bond at the point of cleavage. Although component C2 and Factor B show no apparent N-terminal sequence homology, a limited degree of sequence homology is seen around the sites of proteolytic cleavage.
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PMID:Limited proteolysis of complement components C2 and factor B. Structural analogy and limited sequence homology. 26 15

Limited tryptic digestion of the pyruvate dehydrogenase complex of Escherichia coli or its dihydrolipoyl transacetylase core cleaves the trypsin-sensitive transacetylase subunits into two large fragments, A (lipoyl domain) and D (subunit binding domain). Release of fragments A from the complex does not significantly affect its sedimentation coefficient or its appearance in the electron microscope. Fragment A contains the lipoyl moieties ((3)H-labeled), is acidic with an apparent isoelectric point of about 4.0, has a M(r) of 31,600 as determined by sedimentation equilibrium analysis, and has a swollen or extended structure (f/f(o) = 1.78). Fragment A exhibits anomalous properties, probably due to its acidic nature. It is resistant to staining with Coomassie blue and it migrates on sodium dodecyl sulfate/polyacrylamide gels as if it had a M(r) of 46,000-48,000. Further tryptic digestion converts fragment A into a lipoyl-containing fragment of M(r) 20,000 (fragment B) and eventually into an apparently stable product of estimated M(r) about 10,000 (fragment C). Fragment D has a compact structure of M(r) about 29,600 as determined by sedimentation equilibrium analysis in 6 M guanidinium chloride, and it possesses the intersubunit binding sites of the transacetylase, the binding sites for pyruvate dehydrogenase and dihydrolipoyl dehydrogenase, and the catalytic site for transacetylation. The assemblage of fragments D is responsible for the cube-like appearance of the transacetylase in the electron microscope. High-resolution electron micrographs of the transacetylase show fiber-like extensions, apparently corresponding to tryptic fragment A, surrounding the cube-like core.
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PMID:Subunit structure of dihydrolipoyl transacetylase component of pyruvate dehydrogenase complex from Escherichia coli. 38 41

Anhydrotrypsin, a derivative of bovine trypsin, immobilized on Sepharose tightly adsorbs various peptides containing L-arginine at the carboxyl termini, such as bradykinin and tuftsin. These peptides correspond to the specific products of the action of trypsin-like enzymes. Native trypsin immobilized on Sepharose does not show such strong affinity. Fragment 2, a peptide with 41 amino acid residues, which has been released together with bradykinin from bovine high-molecular-weight kininogen by the action of plasm kallikrein, is also adsorbed on the immobilized anhydrotrypsin. When only the carboxyl-terminal arginine is removed with carboxy-peptidase B, however, the peptide loses its adsorptive ability. Immobilized anhydrochymotrypsin, on the other hand, exerts specific affinity for the peptides which correspond to the products of chymotrypsin. These results suggest that the anhydro-derivatives of serine-proteseas in general may be of great use in the affinity chromatography of respective series of various naturally occurring peptides.
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PMID:Specific isolation of biologically-active peptides by means of immobilized anhydrotrypsin and anhydrochymotrypsin. 57 60

Normal bovine colostral immunoglobulin G1 was subjected to enzymic digestion (pepsin, papain and trypsin) and the resulting fragments separated by a combination of molecularsieve and phosphocellulose chromatography. Fragments F(ab')2 derived from peptic digestion, fragment Fab from papain digestion and fragment Fab(t) from tryptic digestion showed complete antigenic identity with each other. Although fragment F(ab')2 (peptic digestion) had a sedimentation coefficient (S2o,w) of 5.3S, those for fragments Fab' (peptic digestion), Fab (papain digestion) and Fab(t) (tryptic digestion) were found to be 3.9S, 3.7S and 3.7S respectively. The mol.wts. calculated for the various fragments from the sedimentation equilibrium data were: F(ab')2, 104000 +/-200; Fab', 51900+/-340; Fab, 50900+/-230; Fab(t) 50900+/-300. Fragment Fc' (peptic digestion) had an S20,w of 3.2S and a mol. wt. of 42900+/-650; fragment Fc (papain digestion) had an SI0,w of 3.7S and a mol.wt. of 50800+/-300; fragment Fc(t) had an S20,w of 3.7S and a mol.wt. of 50800+/-300; fragment Fc(t) had an S20,w of 3.7S and a mol.wt. of 50800+/-450.
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PMID:Physiochemical characterization of proteolytic cleavage fragments of bovine colostral immunoglobulin G1 (IgG1). 77 68

1. A trypsin and chymotrypsin inhibitor was isolated by extraction of chick-pea meal at pH8.3, followed by (NH4)2SO4 precipitation and successive column chromatography on CM-cellulose and calcium phosphate (hydroxyapatite). 2. The inhibitor was pure by polyacrylamide-gel and cellulose acetate electrophoresis and by isoelectric focusing in polyacrylamide gels. 3. The inhibitor had a molecular weight of approx. 10000 as determined by ultracentrifugation and by polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate. A molecular weight of 8300 was resolved from its amino acid composition. 4. The inhibitor formed complexes with trypsin and chymotrypsin at molar ratios of 1:1. 5. Limited proteolysis of the inhibitor with trypsin at pH3.75 resulted in hydrolysis of a single-Lys-X-bond and in consequent loss of 85% of the trypsin inhibitory activity and 60% of the chymotrypsin inhibitory activity. Limited proteolysis of the inhibitor with chymotrypsin at pH3.75 resulted in hydrolysis of a single-Tyr-X-bond and in consequent loss of 70% of the trypsin inhibitory activity and in complete loss of the chymotrypsin inhibitory activity. 6. Cleavage of the inhibitor with CNBr followed by pepsin and consequent separation of the products on a Bio Gel P-10 column, yielded two active fragments, A and B. Fragment A inhibited trypsin but not chymotrypsin, and fragment B inhibited chymotrypsin but not trypsin. The specific trypsin inhibitory activity, on a molar ratio, of fragment A was twice that of the native inhibitor, suggesting the unmasking of another trypsin inhibitory site as a result of the cleavage. On the other hand, the specific chymotrypsin inhibitory activity of fragment B was about one-half of that of the native inhibitor, indicating the occurrence of a possible conformational change.
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PMID:A trypsin and chymotrypsin inhibitor from chick peas (Cicer arietinum). 79 Dec 69

Cleavage of the two methionine residues in the glycoprotein trypsin inhibitor ovomucoid, variant O1, with CNBr resulted in two fragments whose mol.wts. were approx. 16 600 (fragment LS) and 11 000 (fragment M). Both fragments formed precipitates with antisera to ovomucoid. Fragment LS retained 56% of the trypsin-inhibitory activity of ovomucoid, but fragment M did not inhibit. After reduction and alkylation, the molecular weight of fragment M was unchanged, but fragment LS could be resolved into two segments of peptide chain with mol.wts. of approx. 12000 (fragment L) and 4700 (fragment S). Each of these peptides contained carbohydrate. Marked heterogeneity was observed in the hexose and hexosamine contents of fragment L. This may account for much of the heterogeneity in neutral carbohydrate occurring in ovomucoid preparations. It was found that fragment M was located at the N-terminal end, fragment S was in the centre and fragment L made up the C-terminal portion of the molecule.
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PMID:Active fragments obtained by cyanogen bromide cleavage of ovomucoid. 93 85

The digestion of EF-Tu-GDP (or EF-Tu-GTP) by trypsin [EC 3.4.21.4] under native conditions has been shown to proceed through two different and characteristic stages. 1. In the first phase, the protein is transformed into a fragment (Fragment A) with a molecular weight of 39,000 by exposure to trypsin for a relatively short period of time. Fragment A is unable to catalyze the binding of aminoacyl-tRNA to ribosomes. The ability to promote two partial steps of the binding reaction, i.e., formation of the aminoacyl-tRNA-EF-Tu-GTP ternary complex as well as the methanol-stimulated, ribosome dependent GTPase reaction, was rapidly destroyed. On the other hand, the ability to interact with guanine nucleotides as well as EF-Ts survived well during prolonged digestion. 2. In the second phase of digestion, a nick is introduced in Fragment A to yield two subfragments (Fragments B and C). These two fragments exist as a hybrid molecule which migrates as a single peak on a Sephadex G-75 column, and which dissociates into Fragments B and C only in the presence of 6 M guanidine hydrochloride or 5% sodium dodecyl sulfate. The molecular weights of Fragments B and C, as determined by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, were 22,000 and 12,000 respectively. The hybrid molecule still retained one mole of bound guanine nucleotide and was resistant to further tryptic digestion. 3. Three sulfhydryl groups of EF-Tu were found to be present in Fragment B, both by amino acid analysis of the purified fragments and also by electrophoresis of tryptic digests labeled with N-ethyl[14C]maleimide. 4. The tryptic digestion of EF-Tu-GDP (or EF-Tu-GTP) labeled with N-(1-anilinonaphthyl-4)maleimide (ANM) at SH2 (the second SH), caused a 30% decrease in the fluorescence emission during the first rapid phase of digestion. This indicates that destruction of the hydrophobic environment near SH2 of EF-Tu occurred in the early phase of tryptic digestion. 5. The kinetic studies on the reaction of ANM with EF-Tu before and after tryptic digestion indicated that both Fragment A and the hybrid molecule reacted with ANM in the presence of GTP three to four times more rapidly than in the presence of GDP. Thus, it appears that the ability to induce conformational transition near SH2 by a change of nucleotide ligands is still retained in the hybrid molecule consisting of Fragments B and C.
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PMID:Limited hydrolysis of the polypeptide chain elongation factor Tu by trypsin. Isolation and characterization of the polypeptide fragments. 93 63

The amino-acid sequence of Fragment A from diphtheria toxin is reported. Fragment A (molecular weight, Mr, 21,145) is the major enzymically active fragment produced upon activation of the intact toxin (Mr about 60,000) by limited tryptic digestion and reduction. It, or a similar fragment, is believed responsible for the inhibition of protein synthesis in animal cells exposed to the toxin. Fragment A, which corresponds to the amino terminus of the toxin, is shown here to consist of three major forms (190, 192, and 193 residues) resulting from cleavage by trypsin adjacent to any of three closely spaced arginine residues. All three forms are enzymically active.
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PMID:Amino-acid sequence of fragment A, an enzymically active fragment from diphtheria toxin. 106 Nov 28

1. 'Inhibitor fragment' isolated from human serum albumin degraded by rabbit cathepsin D is composed of one peptide chain with two intrachain disulphide bonds. There are two kinds of inhibitor molecules having different N-terminal amino acids: one is threonine and the other glutamine. 2. Fragment F1, isolated from inhibitor degraded by trypsin, is composed of two chains linked by a disulphide bond. There are three kinds of fragment F1. All have one alpha chain in common, which has an intrachain disulphide bond. They differ by the nature of the chain, which is linked to the alpha chain by a disulphide bond. The epsilon chain is present in trace amounts. The two other chains, beta and gamma, differ by their C-terminal amino acid, which is respectively arginine and lysine. 3. Inhibitor is composed of the last 92 or 89 residues of the human albumin molecule and fragment F1 is composed of two parts of this C-terminal portion of the albumin molecule.
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PMID:Chemical structure of two fragments of human serum albumin and their location in the albumin molecule. 116 60


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