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)

Rat mast cell tryptase was purified to homogeneity from rat tongue by a series of standard chromatographic procedures. Since the enzyme gave band corresponding to molecular mass of 32-35 kDa on sodium dodecyl sulfate polyacrylamide gel electrophoresis and exhibited a molecular mass of 135 kDa on gel filtration, it was presumed to be a noncovalently associated tetramer. The N-terminal amino acid sequence of 50 residues of the enzyme showed the highest degree of homology with the same region in mouse mast cell protease 7 (92%), and less homology to those of tryptases from man and dog, and peritoneal cells of rats and Mongolian gerbils. The inhibitor specificity of rat tongue tryptase was similar to that of rat peritoneal mast cell tryptase free from trypstatin: it was inhibited by alpha 1-antitrypsin, Kunitz-type soybean trypsin inhibitor and Bowman-Birk soybean trypsin inhibitor, but these inhibitors do not inhibit the tryptases from rat skin, human lung, and dog mast cells. Judging from these results, together with other enzymatic properties, the enzyme may be a novel isoform of tryptase in rat tongue. Analysis by differential staining with peroxidase-labeled lectins of the enzyme suggested that it has tri- and/or tetraantennary complex-type oligosaccharides containing a relatively high amount of sialic acid. The immunohistochemical distribution of this enzyme indicated that the reactive antigen was specific in connective tissue but not in mucosal mast cells.
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PMID:Purification and characterization of a novel isoform of mast cell tryptase from rat tongue. 894 53

Inter-alpha-inhibitor (I alpha I) is a serine protease inhibitor present in human plasma. It has a molecular weight of about 220 kDa which encompasses 3 chains including two heavy chains and one light chain. The light chain, known as bikunin, is responsible for the antitryptic activity of I alpha I in the inhibition of various enzymes, such as trypsin and chymotrypsin. Under physiologic or certain pathologic circumstances, several macromolecules related to I alpha I appear in plasma and urine. However, the physiologic role of I alpha I remains unclear. As far as urolithiasis is concerned, two urinary macromolecules related to I alpha I have been isolated and shown to be potent inhibitors of calcium oxalate formation. One of these inhibitors, uronic-acid-rich protein (UAP), has been identified and well characterized. The sequence of the first 18 amino acid residues of UAP is identical with that of bikunin. Furthermore, the immunoreaction between UAP and I alpha I antibody using immunoblot analysis was positive. UAP isolated from the urine of stone formers exhibited less inhibitory activity towards calcium oxalate crystallization than that derived from the urine of healthy subjects. This suggests a structural abnormality of the inhibitor obtained from stone patients. The organic matrix extracted from kidney stones contained a protein antigenically related to I alpha I. We conclude that UAP is a member of I alpha I family taking part in inhibiting calcium oxalate crystallization, and modulating the formation of stones in the urinary tract.
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PMID:Inter-alpha-inhibitor: a protein family involved in the inhibition of calcium oxalate crystallization. 981 21

The inter-alpha-trypsin inhibitor (ITI) family is a group of structurally related plasma serine protease inhibitors. The ITI family members consist of combinations of mature heavy chains named HC1, HC2, HC3 linked to bikunin (a Kunitz-type protease inhibitor) by a covalent interchain protein-glycosaminoglycan-protein cross-link. The biosynthesis of the ITI family members takes place in the liver. In this report we examine the biosynthesis of these proteins using transient transfected COS-7 cells expressing one or more combinations of human ITI chains. The processing and secretion of alpha1-microglobulin and bikunin does not require the ITI heavy chains. A small proportion of the H3 chain seems to be processed into the HC3 form in the absence of the other ITI chains. In contrast, the processing of H2 into HC2 needs the presence of the L chain. The COS-7 cells are able to link the HC2 and HC3 heavy chains with bikunin by means of a chondroitin sulfate bridge, and thus to generate 260-kDa ITI-like proteins as well as pre-alpha-trypsin inhibitor (PalphaI). However, the maturation of the Hl chain into HC1 and the assembly of HC1 inside multichain proteins may take place according to a mechanism which differs from that of the H2 and H3 chains. These results indicate that the assembly of the constituent chains of the ITI-like proteins and PalphaI is not dependent on the liver machinery.
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PMID:Assembly and secretion of recombinant chains of human inter-alpha-trypsin inhibitor in COS-7 cells. 991 30

Two proteins of 17 and 24 kDa, respectively, which were immunologically related to bikunin, were purified from urine of healthy men, using in the last step a trypsin CNBr-sepharose affinity column. These proteins strongly inhibited calcium oxalate (CaOx) crystallization in two in vitro models. In the first model, the presence of 8 microg/ml protein in a medium containing 0.76 mM CaCl2 (with 45Ca) and 0.76 mM ammonium oxalate inhibited the crystallization process by 80%, as estimated by supernatant radioactivity after 60 min of incubation. A similar inhibition was observed in the second turbidimetric model, where the CaOx crystallization kinetics were followed for 10 min at 620 nm in a medium containing 4 mM CaCl2 and 0.5 mM Na2Ox. These proteins were used as standard protein for the development of an enzyme-linked immunosorbent assay (ELISA) in urine. Mean (+/-SEM) urinary bikunin concentration in 18 healthy subjects was 5.01 +/- 0.91 microg/ml. This was a concentration range of strong inhibitory activity in vitro. Bikunin values were nearly 50% lower (2.54 +/- 0.42 microg/ml, P=0.007) in 31 CaOx renal stone formers (having weddelite crystals in their first morning urine) than in the healthy volunteers. A correlation was found between urinary bikunin and alpha-1 microglobulin concentrations in the control group (y=0.73x + 1.09, r2=0.8) while no such correlation existed in the lithiasis group. In conclusion, bikunin exerts a strong inhibitory action of CaOx crystallization in vitro. Its involvement in urinary CaOx crystallization of stone formers is highly probable, based on the significant decrease in its urinary concentration in the majority of stone formers studied.
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PMID:Inhibitory effect of bikunin on calcium oxalate crystallization in vitro and urinary bikunin decrease in renal stone formers. 1009 56

A glycoprotein with a high inhibitory activity against trypsin was isolated in 1961 from human plasma and named inter-alpha trypsin inhibitor (ITI). Since then, several other proteins that share antigenic and structural similarities with ITI have been identified and classified as members of the ITI protein family. These glycoproteins built up from different combinations of four polypeptides HC1, HC2, HC3 and bikunin are encoded by four genes H1, H2, H3, L on three chromosomes. Bikunin has two proteinase inhibitor domains and belongs to the Kunitz-type protease inhibitor family; it displays an inhibitory activity against trypsin, leukocyte elastase and plasmin. The heavy chains do not have any protease inhibitory properties but have the capacity to interact in vitro and in vivo with hyaluronic acid. This binding promotes the stability of the extra-cellular matrix. Consequently, the ITI protein family is suspected of playing a key role in the extra-cellular matrix biology. Isolation of free heavy chains in bronchial secretions and the recent emphasis about the bikunin role in tumoral invasion should enhance the interest about ITI protein family in the pathophysiology of chronic bronchopulmonary diseases or lung cancer progression.
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PMID:[Proteins of the inter-alpha trypsin inhibitor (ITI) family. A major role in the biology of the extracellular matrix]. 1085 62

Two forms of urinary trypsin inhibitor (UTI-1 and UTI-2) were purified from pooled urine of normal male rats to apparent homogeneity by salting out, affinity chromatography, gel filtration, and reverse-phase HPLC. UTIs-1 and 2 were shown to be thermostable glycoproteins with the respective molecular weights of 22,000 and 18,000 estimated by SDS-PAGE. These inhibitors combined with bovine trypsin in a 1:1 molar ratio: the Kd values were 2.5 x 10(-10) and 2.3 x 10(-10) M, respectively. Amino acid composition and sequence analysis indicated that UTI-1 corresponded to rat bikunin of which the amino acid sequence was deduced from a rat liver cDNA clone encoding alpha1-microglobulin [Lindqvist et al. (1992), Biochim. Biophys. Acta 1130, 63-67] except that the protein sequence seemed to lack C-terminal serine, and UTI-2 corresponded to UTI-1 lacking N-terminal 21 amino acid residues.
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PMID:Purification, characterization, and relation to bikunin of rat urinary trypsin inhibitors. 1130 54

Bikunin is a Kunitz-type proteinase inhibitor, which is cross-linked to heavy chains via a chondroitin sulfate chain, forming inter-alpha-inhibitor and related molecules. Rat bikunin was produced by baculovirus-infected insect cells. The protein could be purified with a total yield of 20 mg/liter medium. Unlike naturally occuring bikunin the recombinant protein had no galactosaminoglycan chain. Endoglycosidase digestion also suggested that the recombinant form lacked N-linked oligosaccharides. Bikunin is translated as a part of a precursor, alpha1-microglobulin/bikunin, but the functional significance of the cotranslation is unknown. Our results indicate that the proteinase inhibitory function of bikunin is not regulated by the alpha1-microglobulin-part of the alpha1-microglobulin/bikunin precursor since recombinant bikunin had the same trypsin inhibitory activity as the recombinant precursor. Both free bikunin and the precursor were also functional as a substrate in an in vitro xylosylation system. This demonstrates that the alpha1-microglobulin-part is not necessary for the first step of galactosaminoglycan assembly.
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PMID:Expression of a functional proteinase inhibitor capable of accepting xylose: bikunin. 1136 89

Noninvasive, epitheliochorial placental attachment in the pig is regulated through endometrial production of protease inhibitors. The objective of the present study was to determine if the light-chain serine protease inhibitor of the inter-alpha-trypsin inhibitor family, bikunin, is produced by the porcine endometrium during the estrous cycle and early pregnancy. Western blot analysis revealed the presence of bikunin in uterine flushings of gilts collected during the luteal phase of the estrous cycle and early pregnancy (Days 12-18). However, bikunin unbound to the inter-alpha-trypsin heavy chains was detected only in endometrial explant culture medium obtained from estrus and pregnant (Days 12, 15, and 18) gilts. Endometrial bikunin gene expression was lowest on Day 10 of the estrous cycle and pregnancy, followed by a 30- to 77-fold increase on Day 15 of the estrous cycle and pregnancy. Bikunin gene expression decreased on Day 18 of the estrous cycle, whereas endometrial bikunin gene expression continued to increase in pregnant gilts. Bikunin mRNA was localized to the uterine glands between Days 15 and 18 of the estrous cycle and pregnancy. In addition to its role as a protease inhibitor, bikunin functions in stabilization of the extracellular matrix, which suggests that bikunin could be involved with facilitating placental attachment to the uterine epithelial surface in the pig.
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PMID:Presence of the acute phase protein, bikunin, in the endometrium of gilts during estrous cycle and early pregnancy. 1146 19

Bikunin, an inhibitor of serine proteases, is widely distributed in human tissues, including the skin, and may inhibit tryptase and modulate allergic inflammation. The purpose of the present study was to compare follicular eruptions (FE), so-called atopic skin or perifollicular accentuation, with atopic dermatitis (AD) lesions (ADL) by immunohistochemical analysis using antibodies to bikunin and tryptase. Immunohistochemically, bikunin was colocalized with tryptase in dermal mast cells, and a small quantity of bikunin was also deposited in the intercellular spaces in FE and ADL. The number of bikunin-laden mast cells per 0.78 mm(2) of skin was 78.1+/-7.1 (mean+/-SEM, n=14) in FE, 25.4+/-2.3 (n=10) in normal skin from children and infants, 91.3+/-11.8 (n=10) in ADL, 25.6+/-4.8 (n=5) in nonlesional skin of AD, and 27.8+/-2.0 (n=13) in normal adult skin. The difference between FE and normal control skin from children and infants, between FE and nonlesional skin of AD, and between lesional and nonlesional skin of AD were significant. Based on the above findings and the occasional presence of spongiosis and lymphocyte infiltration, in FE moderate inflammation is apparent histopathologically even though little inflammation is apparent clinically.
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PMID:Quantitative analysis of bikunin-laden mast cells in follicular eruptions and chronic skin lesions of atopic dermatitis. 1252 75

Urinary trypsin inhibitors (uTi) suppress serine proteases during inflammation. After liberation from proinhibitors (P-alpha-I and I-alpha-I) by the white blood cell (WBC) response, uTi readily pass through the kidneys into urine. A key uTi, bikunin, is attached to O-linked and N-linked glycoconjugates. Recently, uTi inhibitors, called uristatins, were found to lack the O-linked glycoconjugates. Monoclonal antibodies were produced using purified uristatin and screened for binding differences to uristatin, bikunin, P-alpha-I, and I-alpha-I. Antibody-binding patterns were characterized using immunoaffinity binding onto protein-chip surfaces and analysis by Surface Enhanced Laser Desorption/Ionization mass spectrometry (SELDI), using specimens from patients and from purified uTi standards. Antibodies were developed and used in an enzyme-linked immunosorbent assay (ELISA) method for uTi measurement in urine and plasma specimens. ELISA was performed on specimens from normal, presumed healthy, controls and from patients who had been screened for inflammation using a high sensitivity C-reactive protein (CRP) test and a complete blood count (CBC). Polyclonal antibody against uTi showed cross-reactivity with the Tamm-Horsfall protein (THP) and with proinhibitors. Screening of anti-uTi monoclonal antibodies (Mab) revealed antibodies that did not cross-react with either of the above, thus providing a tool to measure both uristatin and bikunin in urine with Mab 3G5 and in plasma with Mab 5D11. The monoclonal antibody 5D11 cross-reacts with specific N-linked glycoconjugates of uristatin present in plasma. In ca 96% of healthy adults, uTi were present at <12 mg/l in urine and <4 mg/l in plasma. We also found that patients with an inflammation and a CRP of >2.0 mg/l had higher urinary concentrations of uTi than the control population in every subject. Free uristatin and bikunin pass readily into urine and are primarily bound to heavy chains that constitute the proinhibitor form in plasma.
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PMID:Immunological evaluation of urinary trypsin inhibitors in blood and urine: role of N- & O-linked glycoproteins. 1711 77


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