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 heat-stable protein has been detected in Saccharomyces cerevisiae which inhibits mitochondrial ATPase activity. The protein inhibitor has been isolated from extracts prepared by brief heat treatment of unbroken cell suspensions. The isolated inhibitor is a small basic protein (molecular weight close to 7000, isoelectric proint 9.05) devoid of tryptophan, tyrosine, and cysteine as well as proline. The NHP2-terminal amino acid is serine. The ultraviolet absorption spectrum shows the vibrational fine structure of the phenyl-alanine band. Like the ATPase inhibitor from bovine heart mitochondria the yeast inhibitor is rapidly destroyed by trypsin. It is also inactivated by the yeast proteinases A and B. Radioimmunological analysis indicates that the inhibitor is synthesized on cytoplasmic ribosomes. Its accumulation seems to be connected to the formation of the mitochondrial ATPase complex, since its specific activity is greatly reduced both in extracts obtained from the F1-ATPase-deficient nuclear mutant pet 936 and from the cytoplasmic petite mutant D 273-10B-1.
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PMID:A protein inhibitor of mitochondrial adenosine triphosphatase (F1) from Saccharomyces cerevisiae. 13 3

A noncovalent complex of the apoprotein (1-104) and cyanogen bromide heme fragment containing residues 1 to 65, (1-65) H, has been prepared from horse heart cytochrome c. Conditions under which the redundant portions of the ferrous complex can be removed by limited trypsin digestion have been devised. The complementing fragments have been isolated from the derived complexes and four apofragments and one heme fragment have been identified in the amino acid sequence of cytochrome c. They are (39-104), (40-104), (54-104), (56-104), and (1-53)H. The formation of an ordered ferric complex composed of one heme fragment and one apofragment for the cases (1-53)H (39-104), (1-53)H-(40-104), (1-53)H-(54-104), and (1-53)H-(56-104) has been demonstrated by the quenching of the tryptophan 59 fluorescence and the regain of biological activity in a cytochrome b2 assay. The apparent dissociation constant has been estimated as less than 3 X 10(-7) M in all the aforementioned cases. Thus, the region (between residues 38 and 57) of the amino acid sequence permissible for cleavage without disruption of the ordered structure indicated by the present in vitro experiments corresponds to that (between residues 38 and 57) evolutionally deleted in the three-dimensional structure of Pseudomonas aeruginosa cytochrome c551 discovered by Dickerson et al. (Dickerson, R.E., Timkovich, R., and Almassy, R.J. (1976) J. Mol. Biol. 100, 473-491).
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PMID:Formation of a biologically active, ordered complex from two overlapping fragments of cytochrome c. 19 Feb 31

The exposure of apolipoproteins at the surface of human plasma high density lipoproteins (HDL) was assessed by their accessibility to agarose-immobilized forms of trypsin and chymotrypsin. Proteolysis of lipid-free apolipoproteins and the lipoprotein subfractions HDL2 (d = 1.08--1.125 g/ml) and HDL3 (d = 1.125--1.195 g/ml) that differ in lipid-to-protein ratio was compared by polyacrylamide gel electrophoresis and isoelectric focusing of the apolipoproteins and peptide fragments and by quantitation of the various carboxyl-terminal groups formed. Gel filtration of the proteolyzed lipoproteins on Sephadex G-150 column indicated that more than 90% of the apolipoproteins and peptides remain associated with lipoprotein complexes. Proteolysis of lipoproteins occurred more slowly and with less fragmentation of the lipoproteins and apolipoproteins than proteolysis of thelipid-free apolipoproteins or the proteolysis of lipoproteins by soluble proteases reported by other investigators. The difference in lipid content of HDL2 and HDL3 made little difference in their proteolysis. Proteolysis of the lipoproteins by agarose-trypsin was more rapid at 37 degrees C than at 22 degrees C, but the proteolytic products were similar and differed from the products from the lipid free proteins. Peptide fragments from lipoproteins were larger than those from lipid-free proteins, which suggests masking of potentially cleavable groups by lipid. The amounts (mol/g protein) of new carboxyl-terminal tyrosine and phenylalanine released by agarose -chymotrypsin were much greater from the lipid-free proteins, but about 3/4 of the tryptophan residues were inacessible in both lipoproteins and lipid-free proteins. In agarose-trypsin digestion, lysine residues were slightly more masked than arginine in the absence of lipids and much more so in the lipoproteins. However, in the lipoproteins apoA-II, which contains lysine but no arginine, was cleaved more rapidly and extensively by agarose-trypsin than apoA-I.
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PMID:Surface exposure of apolipoproteins in high density lipoproteins. I. Reactivities with agarose-immobilized proteases. 20 44

Chemical and enzymatic properties of four collagenases newly isolated from anaerobic Clostridium histolyticum, aerobic Achromobacter iophagus, and from two lower eucaryotes, the fungus Entomophthora coronata and the insect Hypoderma lineatum are reviewed. The problems of their biosynthesis and precursors, namely the effect of induction of collagenase and neutral proteinase in Achromobacter by their macromolecular substrates are discussed. The two bacterial collagenases are Zn-metallo-enzymes; the highly purified Clostridium collagenase contains cyst(e)ine, serine phosphate and tryptophan additionally to amino acids reported previously. Achromobacter collagenase has the highest specific activity of all collagenases; it yields by autolysis enzymatically active degraded forms. The active dimer is composed of two identical subunits of molecular weight 35,000. Similarities between Achromobacter collagenase, thermolysin and Bacillus subtilis neutral proteinase in molecular weight, amino acid composition, and amino acids important for the active sites are discussed. The two collagenases from low eucaryotes are serine proteinases; Hypoderma collagenase is homologous to the trypsin family in the amino terminal sequence. The initial cleavage of native collagen by highly purified bacterial collagenases occurs in the central helical part of the alpha chains and not progressively from the amino terminal end. One of the two initial cleavages produced by Achromobacter collagenase is situated in the region cleaved specifically by vertebrate collagenases, but with different bond specificity. The same is true for the insect collagenase. Entomophthora collagenase is a proteinase of broad specificity which also cleaves collagen in its helical parts. All four collagenases also degrade other proteins according to their bond specificity.
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PMID:Some newly characterized collagenases from procaryotes and lower eucaryotes. 22 May 20

The chymotrypsin-like proteins (chymotrypsin-CT,chymotrypsinogen-CTG, trypsin-T and modified chymotrypsins-at Met 192-MCT and at Tyr 146, 171-TCT), gamma-irradiated in the presence of air, were investigated. Irradiation leads to the unfolding of the native structure of CT-like proteins both in solution and in the dry state, which was shown by the tryptophan fluorescence, viscosimetry and microcalorimetry. The radiation yield of unfolded molecules Gconf was estimated and compared with (1) the rate constants for the reactions of OH-radicals with the proteins as determined by the p-nitrosodimethylaniline, (2) general stability of protein globule using the difference of the energies of the unfolded and globular conformations and (3) the radiation yield of tryptophan destruction in proteins-G-trp. There was a correlation between the values of Gconf and G-trp. The ratio G-trp/Gconf, which defines the number of destroyed tryptophan residues for one unfolded protein molecule, was constant within the limits of error. For CT, MCT, TCT and CTG, this ratio was on the average 3-2, and for T it was 2-2 residues. These facts point to the role of tryptophan destruction in the unfolding of the native structure of CT-like proteins on irradiation.
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PMID:Gamma-irradiated chymotrypsin-like proteins. I. Structural changes. 30 Jul 18

The physicochemical and catalytic properties of thioredoxin-T' are described. This complemented protein structure consists of a 1:1 complex between the inactive fragments thioredoxin-T-(1--73) and thioredoxin T-(74--108). These are generated by selective trypsin cleavage at Arg-73 in lysine-modified and denatured Escherichia coli thioredoxin. Thioredoxin-T' was a slowly formed but stable complex with an apparent KD below 10(-8) M. The tryptophan fluorescence spectrum and the CD spectrum were very similar to those of native thioredoxin; some conformational differences were detected by gel chromatography and radioimmunoassay. Thioredoxin-T'-S2 was a substrate for NADPH and thioredoxin reductase and had 1--2% of the activity of native thioredoxin. This low relative activity was the result of a major increase in the Km value. Thioredoxin-(SH)2 was a hydrogen donor for E. coli ribonucleotide reductase with about 3% relative activity. These results for thioredoxin-T' are correlated with the known three-dimensional structure of thioredoxin. The microenvironment around Arg-73 that is close to the active disulfide appears to be of critical importance for the interactions of thioredoxin with thioredoxin reductase and ribonucleotide reductase.
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PMID:Structure and enzymatic functions of thioredoxin refolded by complementation of two tryptic peptide fragments. 39 Dec 70

The 1-dimethylamino-5-naphthalene sulfonyl (dansyl) derivative of alanyllsyl chloromethane was synthesized and employed to introduce the fluorescent dansyl moiety specifically into the active site of trypsin via affinity labelling. The potential of dansylalanyllysyl chloromethane lies in its high degree of selectivity and markedly faster rate of enzyme inactivation when compared to previously synthesized, single residue affinity label chromophores. This permits the practical utilization of stoichiometric amounts of the inhibitor to achieve 100% inactivation of trypsin, even at high dilutions. The transfer of energy between the four tryptophan residues of trypsin and the bound dansyl group has been investigated in the fluorescent inhibitor-enzyme conjugate. From transfer efficiency measurements mean distances of 19.0 A and 19.3 A between the point of attachment of the dansyl group and the four tryptophan residues of trypsin have been calculated. These compare well with the mean value of 18.8 A derived from calculations based on crystallographic model studies.
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PMID:Synthesis, characterization and fluorescence studies on N-alpha-dansylalanyllysyl trypsino (HIS-46)-methane. 42 27

Even at low concentration, phalloidin shows a marked protection of F-actin against the action of trypsin or pronase. G-actin is not protected at any concentration of phallodin. The kinetics of the proteolysis show that a change in the environment of tryptophan residues is preceded by disruption of the filamentous structure of F-actin.
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PMID:Effect of phalloidin on actin proteolysis as measured by viscometry and fluorimetry. 42 29

Transfers of singlet energy within trypsin were investigated by measuring the fluorescence absorption anisotropy of its tryptophan residues. A ratio of the anisotropy of trypsin to that for N-acetyl-L-tryptophanamide was determined between 306 and 250 nm. The ratio had an average value of 0.7, whether the trypsin anisotropy was measured at 228 of 296 K. However, trypsin dissolved in 5 M guanidine hydrochloride showed little fluorescence depolarization at 228 K (the anisotropy ratio was approximately equal to 0.9). Thus, there is an extensive conformation-dependent energy transfer between tryptophans in trypsin. The ratio of anisotropies of tyrpsin at 304--270 nm was used to estimate energy transfer from tyrosine to tryptophan. Ratios of 1.8 and 1.7 were obtained at 296 K for the native and guanidinium-unfolded enzyme, respectively. The comparable value for N-acetyl-L-tryptophanamide was 1.7. This indicates that there is little transfer from tyrosine to tryptophan in trypsin at 296 K. As confirmation, the excitation wavelength dependencies of the indole fluorescence quantum yield were the same for native and unfolded trypsin. When experiments were performed at 228 K, the 304--270-nm anisotropy ratios were 2.6 for native and 2.1 for unfolded trypsin at pH2. This indicates that the efficiency of energy transfer from tyrosine to tryptophan increases at low temperatures. A photochemical source of error in the quantitation of the efficiency of energy transfer from tyrosine to tryptophan is also described.
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PMID:Transfer of singlet energy within trypsin. 47 91

The largest fragment produced by complete cyanogen bromide digestion of the alpha chain of human fibrinogen contains 236 residues and has a calculated molecular weight of 23,949. The complete amino acid sequence of the fragment was determined by the isolation of peptides generated by plasmin, trypsin (including digestion of citraconylated material), staphylococcal protease, and chymotrypsin. In addition, some key subfragmentation was achieved by selective chemical cleavage at tryptophan residues. The fragment has an unusual amino acid composition, more than half of its residues being glycine, serine, threonine, and proline. There are very few nonpolar residues, although 7 of the alpha-chain's 10 tryptophans occur in this fragment. The fragment contains 2 cysteine residues located 30 residues apart which are connected by an intrachain disulfide bond in the native molecule. The tryptophans occur with a definite periodicity that highlights a series of 13-residue homology repeats. The fragment also contains the two principal alpha-chain cross-linking sites.
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PMID:Amino acid sequence studies on the alpha chain of human fibrinogen. Complete sequence of the largest cyanogen bromide fragment. 51 44


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