Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.5.1.4 (deaminase)
 5,113 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Using a direct radioimmunoassay and a kininogenase assay, we determined that 68% of rat urinary kallikrein was enzymatically active while 32% was in an inactive form which was activated by trypsin. Inorganic cations, at concentrations found in rat urine, were inhibitory in an amidase assay but appeared to potentiate kininogenase activity of pure rat urinary kallikrein. In random urines, kinin concentration was 4.2 +/- 0.7 ng/ml. Trypsinization of the urines generated 52.9 +/- 25.8 ng kinin/ml, indicating that kininogen was present. The rate of kinin formation in vivo may depend on the availability of kininogen and the concentration of inorganic cations in urine, as well as on other well-recognized factors, such as the kallikrein activity of the urine.
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PMID:Components of the kallikrein-kinin system in rat urine. 656 18

The collagenolytic serine protease (crab protease) isolated from the hepatopancreas of the fiddler crab, Uca pugilator, has been investigated with respect to its peptide bond specificity and catalytic properties by using noncollagenous substrates. In contrast to vertebrate collagenases, crab protease is a good general protease capable of degrading a variety of polypeptide and synthetic low molecular weight substrates. Crab protease displays a broad range of specificity, cleaving on the carboxyl-terminal side of residues with both positively and negatively charged side chains as well as hydrophobic side chains. The enzyme appears to favor tyrosyl, phenylalanyl, leucyl, and perhaps lysyl residues and, to a lesser extent, arginyl and glutamyl residues. The rate of cleavage of polypeptide substrates is similar to chymotrypsin but is significantly less than trypsin or chymotrypsin for low molecular weight esterase and amidase substrates. Crab protease is effectively inhibited by chymostatin but not by leupeptin or elastatinal. Several common chloromethyl ketone derivatives of phenylalanine and lysine are also ineffective, although crab protease efficiently cleaves at these residues in polypeptide substrates.
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PMID:Substrate specificity of the collagenolytic serine protease from Uca pugilator: studies with noncollagenous substrates. 678 31

Three acidic arginine esterases have been isolated from the venom of the Western Diamondback rattlesnake (Crotalus atrox). These components demonstrated marked differences in ionic characteristics, as noted by KCl gradient elution from a DEAE-Sephadex A-50 column. Molecular weights, as determined from gel permeation chromatography, were estimated at 25,100 (fraction D), 24,000 (B); and 22,900 (F) for the three separate enzymes. The two larger enzymes (B and D) exhibited similar activities toward the synthetic substrates alpha-N-benzoyl-L-arginine ethyl ester and alpha-N-benzoyl-DL-arginine-p-nitroanilide. Hydrolysis rates were similar to commercial trypsin preparations. Fraction F exhibited a markedly lower activity as an arginine esterase and negligible activity as an arginine amidase. Arginine esterase activity was evident for all three enzymes in the presence of ethylenediamine tetra-acetic acid.
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PMID:Purification of three arginine esterases from the venom of the Western Diamondback rattlesnake (Crotalus atrox). 681 57

1. Two trypsin-like enzymes, assayed by their amidase activity with N-alpha-benzoyl-DL-arginine-p-nitroanilide (DL-BAPNA) as the substrate, were isolated from the gut of the arctic fish capelin (Mallotus villosus). 2. Purification involved affinity chromatography (Benzamidine-CH-Sepharose 4B) of the 30 to 70% (NH4)2SO4 precipitation fraction of a crude extract of the gut, followed by DEAE-Sephadex chromatography, yielding two enzymes, designated Enzyme I and II. 3. Both enzymes had MW of about 28,000 as determined by SDS-electrophoresis. Their isoelectric points were 5.6-5.9 (Enzyme I) and 5.1-5.3 (Enzyme II) and they had similar amino acid composition. 4. Both enzymes were inhibited by standard trypsin inhibitors including the serine protease inhibitor phenylmethyl sulphonyl fluoride (PMSF), but not by the chymotrypsin inhibitor L-1-tosylamide-2-phenylethyl chloromethyl ketone (TPCK). 5. The enzymes had a pH optimum of 8-9 and their stability was not affected by CaCl2. Low pH (2.3) caused an initial rapid loss of enzyme activity, followed by relatively slow decomposition of the activity remaining after 1 hr at 4 degrees C. 6. The enzymes had an apparent temperature optimum of 42 degrees C, resulting from rapid self digestion at higher temperatures.
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PMID:Characteristics of two trypsin type isozymes isolated from the arctic fish capelin (Mallotus villosus). 708 13

The objectives of this study were to determine ascorbic acid stability and its effect on antiproteinase activity of seminal plasma in the presence of an oxidant. Effect of seminal plasma, and additives: glutathione, albumin, hydrogen peroxide and Tris buffer, on ascorbic acid degradation was investigated by UV absorbance. Antiproteinase against trypsin amidase activity was measured spectrophotometrically using N-benzoyl-DL-arginine-p-nitroanilide (BAPNA) as substrate. Ascorbic acid was destroyed much more rapidly with the addition of hydrogen peroxide than in Tris buffer at pH 8.2 alone. Seminal plasma protected ascorbic acid more efficiently than glutathione and albumin alone. The protective effect of seminal plasma on ascorbic acid degradation may closely relate to the function of ascorbic acid in reproductive system of scurvy-prone animals including teleost fish. Within the range of 1-8 mM concentrations, ascorbic acid had a pro-oxidant action on seminal plasma antiproteinase activity in vitro when they were incubated with hydrogen peroxide.
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PMID:Protective effect of seminal plasma proteins on the degradation of ascorbic acid. 747 34

Polymyxin acylase isolated from Pseudomonas sp. M-6-3 was used as an N-myristoyl cleaving enzyme in order to determine a part of the N-terminal amino acid sequence of N-myristoyl proteins. The enzyme hydrolyzed a number of N-myristoyl oligopeptides at various hydrolysis rates but not N-myristoyl proteins. The oncogenic protein (N-myristoyl-pp60c-src) was isolated from human colon adenocarcinoma cell line COLO 320DM by two-dimensional polyacrylamide gel electrophoresis. The protein was digested with trypsin and the resultant tryptic N-myristoyl tetrapeptide (N-myristoyl-Gly-Ser-Asn-Lys) was purified by HPLC and the structure was determined both by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI TOF MASS) and by a gas-phase protein sequencer before or after treatment with the polymyxin acylase. The results suggest that the N-myristoyl peptide sequence derived from N-myristoyl proteins was clearly determined by the combined use of MALDI TOF MASS and the N-myristoyl cleaving enzyme.
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PMID:Determination of N-myristoyl peptide sequence both by MALDI TOF MASS and with an N-myristoyl cleaving enzyme (polymyxin acylase). 750 45

alpha-Macroglobulin and murinoglobulin were purified to homogeneity from Syrian hamster plasma and their properties were compared with those of their respective homologs from other mammals. The trypsin-inhibiting capacity of hamster murinoglobulin was much weaker than those of rat and mouse murinoglobulins. Hamster alpha-macroglobulin was cleaved by trypsin at a number of sites whereas the human homolog was split essentially only in a "bait" region into two fragments of similar size. Hamster alpha-macroglobulin treated with methylamine differed from that treated with trypsin in the electrophoretic mobility, intensity of fluorescence induced by binding of bis(8-anilino-1-naphthalenesulfonate), and plasma clearance pattern, whereas virtually no difference was observed between the human homologs treated in the same manner. The reaction of hamster alpha-macroglobulin with methylamine, as measured by the generation of thiol groups and the decrease in trypsin-protein amidase activity, was much slower than that of the human homolog. Trypsin in a complex with hamster alpha-macroglobulin retained its fibrinolytic activity, but this was not the case for human or rabbit alpha-2-macroglobulin. These results suggest that, compared with the human homolog, hamster alpha-macroglobulin is more loosely packed in the native state, undergoes conformational change more slowly on treatment with methylamine, and less efficiently hinders the access of proteinaceous substrates to trapped proteinase. The serum concentration of hamster alpha-macroglobulin was 6.9 mg/ml, or about 3-fold higher than that of the human type, and showed little change during the acute-phase reaction.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Hamster alpha-macroglobulin and murinoglobulin: comparison of chemical and biological properties with homologs from other mammals. 750 51

The glycoprotein bovine fetuin was treated with trypsin and the Asn-81 tryptic glycopeptide was purified (90% pure by Edman sequencing) by reversed-phase chromatography (RP-HPLC). The Asn-81 glycopeptide, which eluted as a single peak by RP-HPLC, was separable into five peaks on the NucleoPac PA100 column, a pellicular anion-exchange column. Each of the five Asn-81 glycopeptide peaks was shown to contain N-linked oligosaccharides by treatment of each peak with peptide N4-(N-acetyl-beta-D-glucosaminyl) asparagine amidase F (PNGase F) and subsequent oligosaccharide analysis by high-pH anion-exchange chromatography with pulsed amperometric detection. High-pH anion-exchange chromatography-pulsed amperometric detection oligosaccharide analysis revealed that each peak contained a different population of sialylated N-linked oligosaccharides. Hence each peak contained a different group of glycopeptide glycoforms. It was observed that the longer the retention time of the Asn-81 glycopeptide peak on the anion-exchange column, the greater the oligosaccharide sialylation. Two glycopeptide peaks which differed in their distribution of disialylated oligosaccharides demonstrated that the glycopeptide separation was a result of something more than gross differences in sialic acid content. The two other N-linked tryptic glycopeptides of fetuin were also separated into multiple peaks on the NucleoPac PA100 column and these separations were shown to be due to differences in oligosaccharide sialylation. The separations of the three fetuin N-linked glycopeptides demonstrate that pellicular anion-exchange chromatography offers improved separation speed and resolution for the separation of sialylated glycopeptides.
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PMID:Improved fractionation of sialylated glycopeptides by pellicular anion-exchange chromatography. 751 57

The mature form of human protein HC, or alpha 1-microglobulin, has been expressed in Escherichia coli. Protein HC is a member of the lipocalin superfamily of hydrophobic ligand-binding proteins, and carries a heterogeneous chromophore linked covalently by a reduction-resistant bond. Protein HC was first overexpressed as a C-LytA/HC fusion protein containing the C-terminal moiety of the pneumococcal lytic amidase (LytA). Recombinant C-LytA/HC was found to be an insoluble aggregate that was solubilized with 6 M guanidinium chloride and renatured by the addition of thiol reagents in the presence of L-arginine. Recombinant protein HC (rHC) was released from C-LytA/HC by trypsin digestion and purified by size-exclusion chromatography. rHC protein possesses an N-terminal amino-acid sequence identical to that of human protein HC, and a slightly lower molecular mass as determined by SDS-PAGE. Both C-LytA/HC and rHC reacted with polyclonal antibodies raised against native protein HC. A photodiode array detection system on-line with a HPLC system has allowed the identification of a chromophore associated to rHC protein displaying significant absorption in the visible region of the spectrum in resemblance to that found in the natural form of human protein HC.
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PMID:Expression of the human complex-forming glycoprotein HC (alpha 1-microglobulin) in Escherichia coli. 753 95

Trypsinogen is a serine protease zymogen (EC.3.4.21.4) which has proved to be of key significance in a family of about 20 structurally and functionally related pancreatic digestive enzymes. This study was an endeavour to isolate, purify and characterize a stable form of ostrich trypsinogen, which has thus far not yet been accomplished. Trypsinogen (anionic) was isolated and purified by alkaline extraction of pancreatic acetone powder, followed by Toyopearl DEAE 650M, hydroxylapatite and LBTI-Sepharose affinity chromatography. The enzyme was chemically physically and kinetically characterized, using amidase and esterase activity and spectrofluorometric determinations. Effects of CaCl2 and pH, among others, were examined. Purification of homogeneous anionic ostrich trypsinogen was achieved. Immunochemical analysis and spectrofluorometric reaction with sulphonyl-Ala-Ala-Pro-Arg-7-amino-4-methylcoumarin indicated trypsin-free ostrich trypsinogen, with an average Mr of 23,016 and a pI of 4.93. N-terminal sequence data revealed an unique activation peptide sequence, VPGDADDDK. Certain concentrations of Ca2+ enhanced trypsinogen activation, whilst others appeared to have the opposite effect. The kcat/Km values obtained at different pHs, using N alpha-benzoyl-DL-arginine-p-nitroanilide, p-toluenesulphonyl-arginine-methylester and p-toluenesulphonyl-lysine-methylester, followed the pH profile activity trend closely, with maximum catalytic activity at about pH 8 for both ostrich and bovine activated trypsinogen. Ostrich trypsin has significantly higher amidase activity than bovine trypsin, while esterase activities of the two enzymes have an inverse ratio. Kinetic pKa values were 7.2 and 7.4 for ostrich and bovine activated trypsinogens, respectively. The existence of ostrich trypsinogen in a now homogeneous stable form, free of autocatalytic inducing impurities, together with its characterization scenario will hopefully make a significant contribution to the field of comparative biochemistry. This study also confirms that ostrich trypsinogen is closely related to its serine protease counterparts.
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PMID:Ostrich trypsinogen: purification, kinetic properties and characterization of the pancreatic enzyme. 764 28


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