Isolation of a product from the trypsin-digested glycoprotein of sciatic nerve myelin.

Biochim Biophys Acta 1978 Sep 26;536(1):122-33   

Roomi MW; Eylar EH

When purified rabbit sciatic nerve myelin , whether lyophilized or not, is treated with low amounts of trypsin (25 microgram/ml) for 0.5, 3 , or 24 h the resulting protein patterns viewed on sodium dodecyl sulfate (SDS) gel electrophoresis are similar. The most striking feature of the trypsinized myelin is the accumulation of a heavy band at the basic protein position, molecular weight 19 000, which is accounted for as a degradation product of the PO protein , referred to as the TPO protein . The PO protein , the major glycoprotein of sciatic nerve myelin , as well as the 23K and P2 proteins and albumin, an absorbed component , are all partially degraded; most high molecular weight bands are lost. The TPO protein , isolated by gel filtration in 2% SDS on an agarose column, like the PO protein , is highly insoluble in aqueous solvents. It is a glycoprotein (8% carbohydrate), staining with periodic acid-Schiff reagent; containing 3 mannose, 1 galactose, 3 N-acetylglucosamine, 1 sialic acid, and 1 fucose residues and is identical to the nonasaccharide of the parent PO protein . The amino acid composition of the TPO protein , is similar to the PO protein , but has a much higher content of hydrophobic residues and begins with NH2-methionine. This suggests that the PO protein is an amphipathic membrane protein in which its more polar character is confined to the first third of its NH2-terminus. This polar domain is probably positioned above the lipid leaflet where it is accessible to trypsin which cleaves a sensitive lysinyl (or argininyl)-methionine linkage. The more hydrophobic domain (the TPO protein ) is buried in the myelin bilayer where it is protected from further tryptic attack. Thus trypsin can serve as a useful probe of myelin structure.