Weak binding and removal of extrinsic proteinase activities of myelin membranes

The concurrent release of myelin basic protein (MBP) and extrinsic proteina ses from isolated myelin membranes by aqueous solvents of high ionic streng th is considered circumstantial evidence of a presumptive mutual interactio n in situ. The joint release of proteins and proteinases from myelin membra nes of bovine brain, depending on the ionic strength of aqueous solvents, w as therefore examined; 25 mM Tris buffer released an average 1.4% of total myelin protein. It was attributable to about 25 different electrophoretic b ands, but no apparent MBP. However, the extract potently mediated the limit ed proteolysis of added MBP at pH 4.0, 5.6, and 9.0. Because of the PH and the effects of specific inhibitors, proteolysis appears to be owing to acti vities of cathepsin B and D, and an alkaline metalloproteinase. The subsequ ent extraction of myelin membranes with buffered 300 mM NaCl released an ad ditional 20% of total myelin protein, mainly MBP. The extracts, unlike thos e of untreated myelin membranes, no longer cleaved MBP at PH 5.6 and 9.0, a nd did so only slightly at pH 4.0. The results indicate that the bulk of so luble myelin-associated proteinases is much less tightly bound than MBP. Th e weak binding of the former and the prevalence of lysosomal cathepsin B- a nd D-like activities suggest that during their isolation, myelin membranes may adsorb soluble cellular proteins of tissue homogenates. At any rate the washing of myelin membranes with dilute buffer was found to largely remove soluble proteinase activities that are otherwise associated with salt-solu ble MBP of myelin.