The effect of pretreatment of human or rabbit platelets with chymotrypsin on their responses to human fibrinogen and aggregating agents

Greenberg, J.; Packham, MA; Guccione, MA; Harfenist, EJ; Orr, JL; Rathbone, RL Kinlough; Perry, DW; Mustard, J. F.

doi:10.1182/blood.v54.4.753.753

Cited by 91 publications

(23 citation statements)

References 39 publications

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“…In contrast, there is little difference between the responses of control and chymotrypsin-treated platelets to the receptor agonist peptide SFLLRN [49]. Because chymotrypsin also cleaves glycoprotein Ib on platelets [50], Kinlough-Rathbone et al [49] proposed two hypotheses to explain the effects of this protease on platelets. Cleavage of glycoprotein Ib by chymotrypsin could remove an interaction that is important for activation of the receptor by thrombin.…”

Section: Cleavage Of the Thrombin Receptor By Pancreatic Serine Protementioning

confidence: 99%

Cleavage of the thrombin receptor: identification of potential activators and inactivators

Parry

Myles

Tschopp

et al. 1996

Biochemical Journal

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The kinetic parameters were determined for the hydrolysis of a peptide based on the activation site of the thrombin receptor (residues 38-60) by thrombin and 12 other proteases. The kcat and Km values for the cleavage of this peptide (TR39-40) by thrombin were 107 s-1 and 1.3 microM; the kcat/Km of TR39-40 is among the highest observed for thrombin. A model is presented that reconciles the parameters for cleavage of the peptide with the concentration dependence of cellular responses to thrombin. Cleavage of TR39-40 was not specific for thrombin. The pancreatic proteases trypsin and chymotrypsin hydrolysed TR39-40 efficiently (kcat/Km > 10(6) M-1.s-1). Whereas trypsin cleaved TR39-40 at the thrombin activation site (Arg41-Ser42), chymotrypsin hydrolysed the peptide after Phe43. This chymotryptic cleavage would result in inactivation of the receptor. The efficient cleavage of TR39-40 by chymotrypsin (kcat/Km approximately 10(6) M-1.s-1) was predominantly due to a low Km value (2.8 microM). The proteases factor Xa, plasmin, plasma kallikrein, activated protein C and granzyme A also hydrolysed TR39-40 at the Arg41-Ser43 bond, but exhibited kcat/Km values that were at least 10(3)-fold lower than that observed with thrombin. Both tissue and urokinase plasminogen activators as well as granzyme B and neutrophil elastase were unable to cleave TR39-60 at appreciable rates. However, neutrophil cathepsin G hydrolysed the receptor peptide after Phe55. Like the chymotryptic cleavage, this cleavage would lead to inactivation of the receptor, but the cathepsin G reaction was markedly less efficient; the kcat/K(m) value was almost four orders of magnitude lower than that for thrombin. In addition to the above cleavage sites, a secondary site for thrombin and other arginine-specific proteases was identified at Arg46, but the cleavage at this site only occurred at very low rates and is unlikely to be significant in vivo.

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Section: Cleavage Of the Thrombin Receptor By Pancreatic Serine Protementioning

confidence: 99%

Cleavage of the thrombin receptor: identification of potential activators and inactivators

Parry

Myles

Tschopp

et al. 1996

Biochemical Journal

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“…In some experiments the platelets were also exposed to chymotrypsin (8 U/ml, Sigma Chemical Co, St. Louis, Mo) for 30 min during the first washing. Platelets exposed to chymotrypsin interact directly with fibrinogen and aggregate without the addition of stimulating agents (18).…”

Section: Normalmentioning

confidence: 99%

Fibrinogen Milano II: A Congenital Dysfibrinogenaemia Associated with Juvenile Arterial and Venous Thrombosis

Haverkate¹,

Koopman²,

Kluft³

et al. 1986

Thromb Haemost

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SummaryA congenitally abnormal fibrinogen was isolated from blood of a young man with deep-vein thrombosis. Two other affected members of his family had three episodes of severe arterial thrombosis. The fibrinogen showed a delayed clotting by thrombin, but a normal clotting by Arvin®, Reptilase®, and prothrom-bin-staphylocoagulase complex. Analysis of the fibrinopeptides A and B by High Performance Liquid Chromatography did not reveal an abnormal peptide structure. The rate of release of A and B peptides by thrombin was strongly delayed, whereas the rate of release of fibrinopeptide A by Arvin® appeared to be normal. The fibrin polymerization rate was normal. Interactions between the abnormal fibrinogen, platelets and the fibrinolytic system were also normal. Evidence is presented that the defective interaction between fibrinogen Milano II and thrombin is associated with a defective binding of thrombin to the fibrin moiety of the abnormal fibrinogen.

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“…Chymotrypsin-treated platelets from the patient aggregated to the same extent as chymotrypsin-treated platelets from controls on exposure to purified fibrinogen (0.4 mg/mL), in the absence of aggregating agents (not shown). 27 Washed normal platelets aggregated to the same extent with all agonists whether they were resuspended in autologous or in patient citrated platelet-poor plasma (PPP). The defect of aggregation of washed platelets from the patient was not corrected by suspending the platelets in citrated PPP from control subjects (not shown).…”

Section: Platelet Aggregation and The Release Reactionmentioning

confidence: 99%

Identification of a new congenital defect of platelet function characterized by severe impairment of platelet responses to adenosine diphosphate

Cattaneo

Lecchi

Randi

et al. 1992

Blood

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This study characterizes a congenital hemorrhagic disorder caused by a platelet function defect with the following features: (1) severely impaired platelet aggregation and fibrinogen or von Willebrand factor (vWF) binding induced by adenosine diphosphate (ADP); (2) defective aggregation, release reaction, and fibrinogen or vWF binding induced by other agonists; (3) normal aggregation and release reaction induced by high concentrations of thrombin or collagen; (4) no further inhibition by ADP scavengers of aggregation, release reaction, and fibrinogen or vWF binding, comparable with those observed for normal platelets in the presence of ADP scavengers; (5) normal membrane glycoprotein (GP) composition and normal binding of the anti-GP Ilb/llla monoclonal antibody 10E5; (6) no acceleration by ADP of ONGENITAL ABNORMALITIES of primary hemothe normal range. Serum thromboxane BZ levels were also normal. The work from Unit 331 in Lyon (analysis ofplatelet glycoproteins) was supported by MGEN and ARC (subvention 6586).

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The effect of pretreatment of human or rabbit platelets with chymotrypsin on their responses to human fibrinogen and aggregating agents

Cited by 91 publications

References 39 publications

Cleavage of the thrombin receptor: identification of potential activators and inactivators

Cleavage of the thrombin receptor: identification of potential activators and inactivators

Fibrinogen Milano II: A Congenital Dysfibrinogenaemia Associated with Juvenile Arterial and Venous Thrombosis

Identification of a new congenital defect of platelet function characterized by severe impairment of platelet responses to adenosine diphosphate

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