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)

Tissue factor pathway inhibitor-2 (TFPI-2), also known as placental protein 5, is a serine protease inhibitor consisting of three tandemly-arranged Kunitz-type protease inhibitor domains. While TFPI-2 is a potent inhibitor of trypsin, plasmin, kallikrein, and factor XIa in the test tube, the function of this inhibitor in vivo remains unclear. In the present study, we investigated the synthesis and secretion of TFPI-2 by cultured endothelial cells derived from human umbilical vein, aorta, saphenous vein, and dermal microvessels to gain insight into its biological function. While all endothelial cells examined synthesized and secreted TFPI-2, dermal microvascular endothelial cells synthesized threefold to sevenfold higher levels of TFPI-2. Approximately 60% to 90% of the TFPI-2 secreted by endothelial cells was directed to the subendothelial extracellular matrix (ECM). When cultured human umbilical vein endothelial cells were stimulated with inflammatory mediators such as phorbol 12-myristate,13-acetate; endotoxin; and tumor necrosis factor-alpha, TFPI-2 synthesis by these cells increased twofold to 14-fold. Recombinant TFPI-2 bound to dermal microvascular endothelial cell monolayers and its ECM in a specific, dose-dependent, and saturable manner with Kd values of 21 and 24 nmol/L, respectively. TFPI-2 interacted with 4.5 X 10(10) sites/cm2 (3 X 10[5] sites/cell) and 2.3 X 10(11) sites/cm2 on endothelial cells and ECM, respectively. In the presence of rabbit anti-TFPI-2 IgG, but not preimmune IgG, endothelial cells dissociated from the culture flask in a time- and IgG concentration-dependent manner. Our findings provide evidence that endothelial cell-derived TFPI-2 is primarily secreted into the abluminal space and presumably plays an important role in maintaining the integrity of the ECM essential for cell attachment.
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PMID:Quantification and characterization of human endothelial cell-derived tissue factor pathway inhibitor-2. 944 54

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

Urinary trypsin inhibitor (UTI), a Kunitz-type protease inhibitor, directly binds to some types of cells via cell-associated UTI-binding proteins (UTI-BPs). Here we report that the 40-kDa protein (UTI-BP(40)) was purified from the cultured human chondrosarcoma cell line HCS-2/8 by UTI affinity chromatography. Purified UTI-BP(40) was digested with trypsin, and the amino acid sequences of the peptide fragments were determined. The sequences of six tryptic fragments of UTI-BP(40) were identical to subsequences present in human link protein (LP). Authentic bovine LP and UTI-BP(40) displayed identical electrophoretic and chromatographic behavior. The UTI-binding properties of UTI-BP(40) and LP were indistinguishable. Direct binding and competition studies strongly demonstrated that the NH(2)-terminal fragment is the UTI-binding part of the LP molecule, that the COOH-terminal UTI fragment (HI-8) failed to bind the NH(2)-terminal subdomain of the LP molecule, and that LP and UTI-BP(40) exhibited significant hyaluronic acid binding. These results demonstrate that UTI-BP(40) is identical to LP and that the NH(2)-terminal domain of UTI is involved in the interaction with the NH(2)-terminal fragment of LP, which is bound to hyaluronic acid in the extracellular matrix.
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PMID:Identity of urinary trypsin inhibitor-binding protein to link protein. 1080 81

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

Urinary trypsin inhibitor (UTI; Mr 40 kDa) is a Kunitz-type protease inhibitor that efficiently inhibits cell-associated trypsin and plasmin activities. The aim of this study is to examine the expression pattern of UTI in the human ovarian carcinoma ascites fluid by Western blotting, zymography, immunoprecipitation, immunohistochemistry, biochemical and gene analyses and animal experiments. We have identified and characterized the 40 kDa immunoreactive UTI (UTI(40)) and 8 kDa degradation fragment (UTI(8)) in ascites fluid. The levels of UTI(40) and UTI(8) are elevated in ascites fluid taken from patients with ovarian carcinoma relative to paired plasma samples. The UTI(40) and UTI(8) were identified immunologically by the reactivity with 2 different anti-UTI antibodies recognizing different epitopes of the UTI molecule, functionally by its ability to bind trypsin and structurally by its apparent molecular mass with and without deglycosylation treatment. The purified polypeptides have been sequenced and were identical with sequences obtained from UTI and the carboxyl-terminal domain of UTI, respectively. However, UTI mRNA was not detected in the ovarian carcinoma tissue and ovarian carcinoma cell lines examined. Based on extravasation experiments using intravenously injected biotinylated inter-alpha-trypsin inhibitor (IalphaI; a precursor of UTI), we conclude that UTI(40) and UTI(8) found in the ascites fluid may result from (i) the extravasation of plasma proteins such as IalphaI into the peritoneal cavity via hyperpermeable vessels and (ii) the subsequent degradation of IalphaI and UTI(40) by tumor cell-associated trypsin-like enzymes.
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PMID:Identification and characterization of a Kunitz-type protease inhibitor in ascites fluid from patients with ovarian carcinoma. 1086 51

Protease inhibitors from potato juice of cv. Elkana were purified and quantified. The protease inhibitors represent ca. 50% of the total soluble proteins in potato juice. The protease inhibitors were classified into seven different families: potato inhibitor I (PI-1), potato inhibitor II (PI-2), potato cysteine protease inhibitor (PCPI), potato aspartate protease inhibitor (PAPI), potato Kunitz-type protease inhibitor (PKPI), potato carboxypeptidase inhibitor (PCI), and "other serine protease inhibitors". The most abundant families were the PI-2 and PCPI families, representing 22 and 12% of all proteins in potato juice, respectively. Potato protease inhibitors show a broad spectrum of enzyme inhibition. All the families (except PCI) inhibited trypsin and/or chymotrypsin. PI-2 isoforms exhibit 82 and 50% of the total trypsin and chymotrypsin inhibiting activity, respectively. A strong variation within the latter activities was shown within one family and between protease inhibitor families.
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PMID:Relative abundance and inhibitory distribution of protease inhibitors in potato juice from cv. Elkana. 1140 80

Hepatocyte growth factor activator inhibitor type-2/placental bikunin (HAI-2/PB) is a serine proteinase inhibitor that contains 2 Kunitz-domains and a presumed transmembrane domain. It has broad inhibitory spectra against various serine proteinases showing potent inhibitory activities not only to hepatocyte growth factor activator but also to plasmin, trypsin and kallikreins. In this study, we investigated the expression of HAI-2/PB in human gliomas in vivo and the effects of HAI-2/PB on the fibrinolytic and invasive capabilities of human glioblastoma cells in vitro. With RNA blot analysis, HAI-2/PB mRNA was expressed in normal brain and in low-grade astrocytomas, but was hardly detectable in anaplastic astrocytomas and glioblastomas, indicating that its expression levels were inversely correlated with the histological grade of human gliomas. To further explore the possible role of HAI-2/PB in glioma progression, cultured human glioblastoma cell lines (U251 and YKG-1) were transiently transfected with an expression vector harboring human HAI-2/PB cDNA. Subsequent analysis indicated that the expression of HAI-2/PB suppressed the fibrinolytic activities of both glioblastoma cell lines. Moreover, HAI-2/PB inhibited Matrigel invasion of U251 and YKG-1 cells by 30% and 64%, respectively. This anti-invasive effect appeared to be mediated primarily by the inhibitory activity of HAI-2/PB against the serine proteinase-dependent matrix degradation. These findings suggest that the reduced expression of HAI-2/PB is possibly involved in the progression of human gliomas.
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PMID:Reduced expression of hepatocyte growth factor activator inhibitor type-2/placental bikunin (HAI-2/PB) in human glioblastomas: implication for anti-invasive role of HAI-2/PB in glioblastoma cells. 1143 97

Recently we have described a novel secreted protein (the WFIKKN protein) that consists of multiple types of protease inhibitory modules, including two tandem Kunitz-type protease inhibitor-domains. On the basis of its homologies we have suggested that the WFIKKN protein is a multivalent protease inhibitor that may control the action of different proteases. In the present work we have expressed the second Kunitz-type protease inhibitor domain of the human protein WFIKKN in Escherichia coli, purified it by affinity chromatography on trypsin-Sepharose and its structure was characterized by CD spectroscopy. The recombinant protein was found to inhibit trypsin (Ki = 9.6 nm), but chymotrypsin, elastase, plasmin, pancreatic kallikrein, lung tryptase, plasma kallikrein, thrombin, urokinase or tissue plasminogen activator were not inhibited by the recombinant protein even at 1 microm concentration. In view of the marked trypsin-specificity of the inhibitor it is suggested that its physiological target may be trypsin.
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PMID:Expression, purification and characterization of the second Kunitz-type protease inhibitor domain of the human WFIKKN protein. 1270 70

Five Kunitz protease inhibitor group B genes were isolated from the genome of the diploid non-tuber-forming potato species Solanum palustre. Three of five new genes share 99% identity to the published KPI-B genes from various cultivated potato accessions, while others exhibit 96% identity. Spls-KPI-B2 and Spls-KPI-B4 proteins contain unique substitutions of the most conserved residues usually involved to trypsin and chymotrypsin-specific binding sites of Kunitz-type protease inhibitor (KPI)-B, respectively. To test the inhibition of trypsin and chymotrypsin by Spls-KPI proteins, five of them were produced in E. coli purified using a Ni-sepharose resin and ion-exchange chromatography. All recombinant Spls-KPI-B inhibited trypsin; K(i) values ranged from 84.8 (Spls-KPI-B4), 345.5 (Spls-KPI-B1), and 1310.6 nM (Spls-KPI-B2) to 3883.5 (Spls-KPI-B5) and 8370 nM (Spls-KPI-B3). In addition, Spls-KPI-B1 and Spls-KPI-B4 inhibited chymotrypsin. These data suggest that regardless of substitutions of key active-center residues both Spls-KPI-B4 and Spls-KPI-B1 are functional trypsin-chymotrypsin inhibitors.
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PMID:Kunitz-type protease inhibitors group B from Solanum palustre. 1763 28

SKPI (shrimp Kunitz-type protease inhibitor) from Marsupenaeus japonicus is a member of serine protease inhibitors which play an important role in the arthropod immunity. To fully understand its function in the innate immunity of shrimp, the skpi gene was cloned into a modified pPIC9K vector with a 6-His tag and expressed by Pichia pastoris GS115. The secretory SKPI was purified from the medium with high purity by using Ni Sepharose High Performance. This results also indicated that the purified SKPI could inhibit the activity of trypsin specifically.
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PMID:[Expression of a shrimp Kunitz-type protease inhibitor in Pichia pastoris and activity analysis]. 1858 30


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