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Cleavage site of calcium-dependent protease in human platelet membrane glycoprotein Ib

K Yamamoto, G Kosaki, K Suzuki

    Thrombosis Research
    |July 1, 1986
    PubMed

    Insights

    Calcium-dependent proteases cleave glycoprotein Ib alpha-chain (GPIb alpha) into distinct fragments. Endogenous proteases release glycocalicin from human platelets, highlighting protease roles in platelet glycoprotein processing.

    Area of Science:

    • Biochemistry
    • Molecular Biology
    • Hematology

    Background:

    • Glycoprotein Ib (GPIb) is a key receptor on human platelets.
    • Calcium-dependent proteases play roles in cellular processes.
    • Understanding GPIb alpha cleavage is crucial for platelet function research.

    Purpose of the Study:

    • To investigate the cleavage sites of glycoprotein Ib alpha-chain (GPIb alpha) by calcium-dependent proteases.
    • To differentiate the actions of exogenous and endogenous proteases on GPIb alpha.
    • To identify the fragments produced during GPIb alpha hydrolysis.

    Main Methods:

    • Enzymatic digestion of purified GPIb alpha and partially purified GPIb (alpha beta-dimer) with m-type and mu-type proteases.
    • Inhibition studies using protease inhibitors (E-64-C, leupeptin) and calcium chelation.
    • Two-dimensional gel electrophoresis to analyze protein fragments.
    • Stimulation of surface-labeled platelets with A23187 to activate endogenous proteases.

    Main Results:

    • Exogenous m-type calcium-dependent protease cleaved GPIb alpha into 100 kDa and 38 kDa fragments.
    • Both m-type and mu-type proteases produced a 100 kDa fragment from GPIb.
    • Endogenous calcium-dependent protease activation released glycocalicin and glycoprotein V from platelet membranes.
    • Released glycocalicin was further digested to a 100 kDa fragment by m-type protease.

    Conclusions:

    • GPIb alpha is hydrolyzed at two sites by exogenous calcium-dependent proteases, yielding glycocalicin and a 100 kDa fragment.
    • Endogenous platelet proteases cleave GPIb alpha at one site, releasing glycocalicin.
    • These findings elucidate the specific cleavage patterns of GPIb alpha by different calcium-dependent proteases.

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