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Identification and characterization of the cytoplasmic antiproteinase (cap) in human platelets - evidence for the interaction of cap with endogenous platelet proteins

Academic Article
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Overview

authors

  • Riewald, Matthias
  • Morgenstern, K. A.
  • Schleef, R. R.

publication date

  • 1996

journal

  • Journal of Biological Chemistry  Journal

abstract

  • To define the presence and potential role of platelet-associated protease inhibitors, we initiated a study designed to characterize the platelet components that are responsible for the formation of two SDS-stable complexes of approximately 58 and 70 kDa initially observed following the incubation of 125I-thrombin and human platelets. We demonstrate that thermal-mediated unfolding of the 58-kDa complex between 125I-thrombin and a nonsecreted platelet protein leads to an apparent molecular mass of 70 kDa. This platelet component is functionally and immunologically indistinguishable from the cytoplasmic antiproteinase (CAP), also known as placental thrombin inhibitor, a recently cloned member of the ovalbumin family of intracellular serpins (serine proteinase inhibitors). CAP-specific mRNA and antigen were detected in human platelets, suggesting that CAP synthesis occurs concurrent with platelet development. Utilizing quantitative immunoblotting, CAP antigen was estimated at 1.014 +/- 0.181 microg/10(9) nonstimulated platelets. After platelet activation with the calcium ionophore A23187, CAP antigen was detected in released microparticles at approximately 0. 195 +/- 0.031 microg/10(9) platelets and a fraction of platelet CAP was proteolytically modified. We provide evidence that these lower molecular mass species arise by cleavage of CAP at or near the reactive site loop. Most importantly, molecular sieving chromatography indicates the presence of an approximately 68-kDa SDS-labile complex between cleaved CAP and a cellular component in A23187-stimulated platelets, suggesting a physiological target of this intracellular serpin and a potential role for this inhibitor in regulating proteolytic activity that may be formed during platelet activation.

subject areas

  • Base Sequence
  • Blood Platelets
  • Blood Proteins
  • Blotting, Western
  • Calcimycin
  • DNA Primers
  • Hydrolysis
  • Iodine Radioisotopes
  • Molecular Sequence Data
  • Platelet Activation
  • Proteins
  • Sodium Dodecyl Sulfate
  • Thrombin
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Identity

International Standard Serial Number (ISSN)

  • 0021-9258

PubMed ID

  • 8636153
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Additional Document Info

start page

  • 7160

end page

  • 7167

volume

  • 271

issue

  • 12

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