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Basic Protein Dissociating from Myelin Membranes at Physiological Ionic Strength and pH Is Cleaved into Three Major Fragments
Author(s) -
Glynn Paul,
Chantry Andrew,
Groome Nigel,
Cuzner M. Louise
Publication year - 1987
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1987.tb05581.x
Subject(s) - deamidation , myelin basic protein , chemistry , membrane , myelin , biochemistry , ionic strength , biophysics , proteolysis , peptide , proteases , extracellular , membrane protein , enzyme , biology , neuroscience , aqueous solution , central nervous system
Experiments were performed with isolated human myelin membrane preparations to analyse factors that may modulate association of myelin basic protein (MBP) with the membranes and could contribute to demyelinating processes. Transfer of membranes (5 mg protein ml −1 ) at 37°C and pH 7.4 from a hypotonic medium, in which they were relatively stable, to one of physiological ionic strength produced three major effects: (1) initial dissociation of MBP from the membranes by a nonenzymatic process that was doubled in the presence of millimolar Ca 2+ /Mg 2+ ; (2) within 10 min, the appearance in the medium of three major MBP fragments (14.4, 10.3, and 8.4 kilodaltons); and (3) progressive acidification of dissociated MBP and its fragments, probably due to deamidation. Between 1 and 6 h a steady state was apparent in which protein equivalent to 15% of the MBP originally bound to the membranes was found in the medium. The three major MBP fragments formed two thirds of this solubilised material and appeared metabolically stable for 24 h. The kinetics of peptide formation suggested that dissociated, rather than membrane‐bound, MBP was cleaved by myelin‐associated neutral proteases. Two‐dimensional electrophoresis and immunoblotting using two monoclonal antibodies indicated that proteolysis occurred in the vicinity of residues 35 and 75. Evidence was also obtained for removal of C‐terminal arginines and relatively rapid deamidation in the C‐terminal half of MBP. These modifications of MBP might also occur if extracellular fluid gained access to the compacted cytoplasmic space of the myelin sheath.