Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.4 (trypsin)
 42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The heterodimeric vitronectin receptor (VNR) and platelet glycoprotein IIb/IIIa (GPIIb/IIIa) are two members of the integrin family of cell adhesion receptors that share the same beta subunit (GPIIIa). These proteins are involved in binding to vitronectin, fibrinogen and fibronectin and in cytoskeleton-membrane interactions. The present study shows that the human placental syncytiotrophoblast brush border membrane contains a heterodimer of subunit Mr values of 140,000 and 90,000 (non-reduced) or 125,000 and 100,000 (reduced). This protein was recognized by a monoclonal antibody to GPIIIa, rabbit antisera to the VNR and a human alloantiserum to GPIIIa. Brush border VNR-related protein bound to an immobilized peptide containing the Arg-Gly-Asp sequence and, less avidly, to immobilized fibrinogen. Only a small fraction of brush border VNR was associated with a cytoskeleton fraction. Membrane-bound brush border GPIIIa was distinct from that of platelets in its resistance to digestion by trypsin and Staphylococcus aureus V8 protease, and had a slightly lower mobility on SDS/PAGE. In addition, lectin-binding studies indicate glycosylation differences between microvillar and platelet GPIIIa heterodimers. Thus, although placental syncytiotrophoblast expresses a beta 3 integrin in its apical brush border, differences in protease sensitivity and carbohydrate content suggest that it may lack or mask certain antigenic determinants. This may be beneficial in avoiding harmful maternal alloantibody responses during pregnancy. Immunohistology showed that the VNR was present in syncytiotrophoblast apical but not basal plasma membranes, and was absent from other forms of trophoblast. The brush border VNR could function in localizing Arg-Gly-Asp-sequence-containing plasma proteins to the materno-trophoblastic interface.
 ...
PMID:A vitronectin-receptor-related molecule in human placental brush border membranes. 172 Jun 17

Albolabrin is a 73 amino acid peptide isolated from the venom of Trimeresurus albolabris. It contains an RGD sequence and 12 cysteines and is a potent inhibitor of both platelet aggregation and fibrinogen binding to the GPIIb/IIIa complex. This protein shows a high degree of analogy (primarily due to the alignment of all cysteines and the RGD) with a number of other viper venom proteins which inhibit cell adhesion and platelet aggregation and are referred to as disintegrins: rhodostomin, trigramin, flavoridin, applagin, elegantin, and batroxostatin. In this study, we found that the reduction and vinylpyridylethylation of albolabrin and flavoridin decreased their platelet aggregation inhibitory activity approximately 40-50 times. It can be postulated that the higher potency of native and reduced flavoridin as compared to albolabrin depends on the substitution of the Asp of albolabrin with a Phe at the C-terminal end of the RGD in flavoridin. The activity of a synthetic C-terminal peptide derived from flavoridin (residues 35-65) containing four cysteines was about 75 times lower than that of the original flavoridin. The substitution of a pair of cysteine residues with alanines in this peptide resulted in further loss of activity. In order to identify the disulfide bonds in albolabrin, the molecule was digested consecutively by trypsin and porcine pancreatic elastase. Peptides resulting from this digestion were isolated by reverse-phase HPLC and identified by amino acid composition and mass spectrometry.(ABSTRACT TRUNCATED AT 250 WORDS)
 ...
PMID:Identification of the disulfide bond pattern in albolabrin, an RGD-containing peptide from the venom of Trimeresurus albolabris: significance for the expression of platelet aggregation inhibitory activity. 203 89

We have obtained evidence that the ligand-recognition region of the integrin beta-subunit, platelet glycoprotein IIIa (GPIIIa), is discontinuous. Receptor function can be localized to residues near the N-terminus and to the central region of the polypeptide chain. The epitope recognized by our monoclonal antibody, CS-1, which substantially inhibits fibrin(ogen) binding to ADP- and thrombin-stimulated platelets [Ramsamooj, Doellgast & Hantgan (1990) Thromb. Res. 58, 577-592], is contained within residues 349-422 of GPIIIa. This sequence is adjacent to a proteinase-resistant domain of GPIIIa which is linked by disulphide bond(s) to an N-terminal segment near to the putative Arg-Gly-Asp recognition site [D'Souza, Ginsberg, Burke, Lam & Plow (1988) Science 242, 91-93]. Limited trypsin digestion of purified platelet GPIIIa yielded a mixture of two-chain molecules comprised of an N-terminal fragment disulphide-bonded to one of four fragments, which began at residues 299, 303, 353 or 423. Tryptic cleavage of the 300-422 segment correlated with loss of immunoreactivity with anti-GPIIIa monoclonal antibody, CS-1. Chymotrypsin cleavage of GPIIIa resulted in an N-terminal 19 kDa fragment joined by at least one intrachain cystine residue to a 46 kDa polypeptide beginning at residue 349. Partial reduction with dithiothreitol released the larger chymotryptic fragment with its epitope for CS-1 intact. These results have enabled us to localize the epitope recognized by our inhibitory monoclonal antibody, CS-1, to residues 349-422 of GPIIIa. Our data are consistent with a structure in which both the N-terminal and central regions of GPIIIa, which may be in close proximity in the functional GPIIb-IIIa complex, participate in ligand binding.
 ...
PMID:Evidence that the central region of glycoprotein IIIa participates in integrin receptor function. 206 10

Spontaneous platelet aggregation appeared in a patient with von Willebrand disease type IIB during the 37th week of pregnancy. This phenomenon was not associated with symptoms of thrombosis and the patient delivered by caesarean section with no complications. Her platelet-poor plasma (PPP) aggregated normal platelet-rich plasma (PRP) and washed platelets. Aggregation was inhibited by monoclonal antibodies with known specificity for the platelet receptors of von Willebrand factor (vWF), i.e. the glycoprotein Ib (GPIb) and the GPIIb/IIIa complex. A monoclonal antibody, which selectively inhibits the binding of vWF to the GPIIb/IIIa complex, did not block aggregation, suggesting that spontaneous aggregation is not dependent on the binding to GPIIb/IIIa of vWF from patient plasma. Aggregation induced by patient plasma could also be blocked either by two monoclonal antibodies raised against vWF or by a fragment derived from trypsin digestion of normal vWF which blocks the ristocetin-induced binding of normal vWF to platelets. These findings indicate that the spontaneous platelet aggregation in this patient results from the binding of her vWF to GPIb but is independent from the binding of her vWF to GPIIb/IIIa.
 ...
PMID:Spontaneous platelet aggregation during pregnancy in a patient with von Willebrand disease type IIB can be blocked by monoclonal antibodies to both platelet glycoproteins Ib and IIb/IIIa. 237 28

A rat monoclonal IgG2a antibody, 5G11, was raised against native human platelet thrombospondin (TSP). Western blot analysis revealed that 5G11 bound (i) to TSP before and after disulfide reduction, and (ii) to a 15-kDa fragment released after prolonged trypsin digestion. Crossed immunoelectrophoresis confirmed that the binding epitope was expressed in the presence of Ca2+ and after treatment of TSP with EDTA. Since 5G11 had no effect on platelet aggregation, the antibody was used to immunoprecipitate Ca2+-dependent and Ca2+-independent TSP-binding molecules on the surface of thrombin-activated surface-labeled 125I-platelets. The experimental basis was that ligand-receptor interactions are of high affinity and that anti-ligand antibodies should precipitate the ligand-receptor complex. With platelets activated in the presence of EDTA, 5G11 predominantly precipitated a 125I-labeled band of Mr 88,000, identified as glycoprotein (GP) IV. In contrast, in the presence of 2 mM Ca2+ and 1 mM Mg2+, 5G11 precipitated a complex of five radiolabeled proteins, among which GPIIb, GPIIIa and GPIV were the most prominent.
 ...
PMID:Identification of platelet membrane thrombospondin binding molecules using an anti-thrombospondin antibody. 246 42

The proteolytic digestion of GPIIIa on intact platelets by chymotrypsin, thrombin, plasmin, trypsin, and staphylococcal V8 protease was monitored in immunoblot studies employing three different antibodies to GPIIIa, one of which was made against a 13-residue synthetic peptide containing the amino terminus of GPIIIa. Chymotrypsin, plasmin, and trypsin gave similar patterns, from which it could be inferred that the major products after extensive digestion were two-chain molecules composed of amino-terminal fragments of Mr approximately 17,000-18,000 disulfide bonded to carboxyl-terminal remnants of Mr approximately 58,000-70,000. These patterns suggest that GPIIIa contains a large disulfide-bonded loop of at least 325 amino acids that is susceptible to proteolytic cleavage, and that the 4 cysteine residues between residues 177 and 273 bond with each other. Such a structure can also account for the presence of the PIA1 epitope, which has recently been localized to a polymorphism at position 33 on these late digestion products. Thrombin did not proteolyze GPIIIa even at 2.5 units/ml. Still to be resolved is whether the minor immunoreactive GPIIIa bands found on normal platelets are related to in vivo or in vitro proteolysis and whether GPIIIa proteolysis plays a role in chymotrypsin-induced exposure of the GPIIb/IIIa receptor.
 ...
PMID:Evidence that platelet glycoprotein IIIa has a large disulfide-bonded loop that is susceptible to proteolytic cleavage. 252 61

Involvement of platelet membrane glycoproteins (GP) in interactions between platelets and tumor cells was studied by using two human tumor cell lines and two monoclonal antibodies against platelet membrane GP. HMV-I cells derived from vaginal melanoma induced platelet aggregation in heparinized plasma, which was not followed by coagulation. M7609 cells derived from colon adenocarcinoma also induced platelet aggregation in heparinized plasma, which, on the contrary, was followed by coagulation. Aggregating activities of the HMV-I cells were abolished by pretreatment with neuraminidase or trypsin, but M7609 activity was not labile to these enzymes. Aggregations induced by M7609 were inhibited by hirudin or MD805, while those by HMV-I were not. M7609 cells dose dependently shortened the recalcification time of normal as well as Factor IX-deficient plasmas, while they were not effective in shortening the time of Factor II- or Factor VII-deficient plasmas. The procoagulant activity of HMV-I cells was 1000 times less than M7609 on the basis of cell numbers. When human platelets were preincubated with monoclonal anti-GPIb or anti-GPIIb/IIIa complex antibodies, neither cell line could cause aggregations. These findings suggest that both GPIb and the GPIIb/IIIa complex on the platelet surface are involved in the thrombin-dependent and -independent platelet aggregations induced by tumor cells.
 ...
PMID:Involvement of platelet membrane glycoprotein Ib and glycoprotein IIb/IIIa complex in thrombin-dependent and -independent platelet aggregations induced by tumor cells. 291 Apr 73

Mn2+ at low (microM) concentrations fulfils the divalent cation requirement for spreading of BHK21 cells on fibronectin. At much higher concentrations, Mn2+ (and to a small extent also Mg2+) induces spreading on haemoglobin, not normally an adhesive protein. Since high Mn2+ also induces spreading of BHK variants unresponsive to exogenous fibronectin, it is unlikely to be acting as a cofactor for secreted cellular fibronectin or by stimulating its secretion. High Ca2+, but not Mg2+, inhibits the induction of spreading by Mn2+ on haemoglobin. Pre-treatment of cells with high concentrations of trypsin decreases the rate of spreading induced by Mg2+ on fibronectin, and by Mn2+ on haemoglobin, to similar extents. High and low Mn2+ could induce spreading, either by different mechanisms or through a common pathway. In the second case, at both concentrations, Mn2+ could act by binding to Ca2+/Mg2+ sites in one or more receptors for adhesion proteins. This would require binding of Mn2+ or Mg2+ to these sites to activate the receptors in the absence of adhesion proteins, and the effect of such proteins to be to increase the affinity of the sites for metal ions. The sites in question may be formed by sequences homologous to those found in the extracellular domains of the vitronectin receptor and platelet membrane glycoprotein IIb. Although very similar to the Ca2+-binding loop in the EF hand of calmodulin, these sequences more closely resemble bacterial galactose-binding protein in lacking one of the conserved co-ordinating side-chains.
 ...
PMID:Induction of fibroblast spreading by Mn2+: a possible role for unusual binding sites for divalent cations in receptors for proteins containing Arg-Gly-Asp. 319 4

The mechanism of association of the human platelet membrane GPIIb-GPIIIa-Ca2+ complex was studied by treating solubilized membranes with various enzymes and cationic peptides and by studying the binding of 45Ca2+ and 125I-fibrinogen before and after dissociation with EGTA and association with Ca2+. Neuraminidase shifted the complex cathodally (presumably due to cleavage of negatively charged domains), whereas trypsin had no such effect. The EGTA-dissociated complex was almost completely reassociated with neuraminidase or the cationic peptide, tetralysine. The monoclonal antibody 10E5, which specifically binds to the Ca2+-associated complex (not to its dissociated components), also bound to the neuraminidase-associated complex. Thus, Ca2+ is not necessary for the association of the complex. Neuraminidase treatment of washed intact platelets resulted in a cathodal shift of the membrane Triton X-100-extracted associated complex with no effect on its ability to dissociate in the presence of EGTA. Neuraminidase treatment of ADP-perturbed washed platelets also resulted in a cathodal shift of the associated complex; however, dissociation with EGTA was inhibited. Thus, critical neuraminidase-sensitive components of the complex (sialic acid residues) are not exposed on the surface of the platelet membrane of resting platelets, but do become accessible following platelet stimulation with ADP or membrane solubilization with Triton X-100. 45Ca2+ bound to the associated complex, to GPIIb of the dissociated complex (not to GPIIIa), to the Ca2+-reassociated complex, and to the neuraminidase-associated complex which had been dissociated with EGTA. Thus, neuraminidase-sensitive components of the solubilized membrane are not required for Ca2+ binding. 125I-fibrinogen bound to the associated complex (not the dissociated complex), to the Ca2+-reassociated complex, and to the neuraminidase-reassociated complex which had been dissociated with EGTA. Thus, Ca2+ is not necessary for 125I-fibrinogen binding to the major antigen complex.
 ...
PMID:Crossed immunoelectrophoresis of human platelet membranes. Effect of charge on association and dissociation of the glycoprotein GPIIb-GPIIIa membrane complex. 377 33

We have previously demonstrated the isolation of platelet membrane glycoprotein IIb-IIIa by affinity chromatography with a specific monoclonal antibody. We have now separated the polypeptide subunits IIb and IIIa of the isolated glycoprotein by preparative sodium dodecyl sulfate polyacrylamide gel electrophoresis and have compared their structural features. Both IIb and IIIa contain approximately 15% carbohydrate, but IIIa contains a larger percentage of mannose residues, suggesting the presence of high mannose as well as complex N-linked oligosaccharide chains. The amino acid compositions are sufficiently similar to imply areas of sequence homology between the two subunits. To examine further the relationship between the subunits, we digested a mixture of 125I-IIb and 131I-IIIa with trypsin and then separated the radiolabeled peptides by high performance liquid chromatography. The resultant peptide maps of IIb and IIIa are completely different. This indicates that neither subunit is derived from the other and suggests that polypeptides IIb and IIIa are products of separate genes.
 ...
PMID:Isolation and structural characterization of the polypeptide subunits of membrane glycoprotein IIb-IIIa from human platelets. 705 67


1 2 Next >>