Incorporation of P0 protein into liposomes: demonstration of a two-domain structure by immunochemical and PAGE analysis.
The amphiphilic nature of P0, the major glycoprotein of peripheral nerve myelin, has been suggested previously. In the present study, purified P0 from human peripheral nerve myelin was incorporated into an artificial lipid bilayer consisting of dimyristoyl lecithin and cholesterol. The liposomes were fractionated on a sucrose gradient. The continued expression of P0 antigenicity by the liposomes was shown by specific complement consumption with a multivalent antiserum against P0 or with an IgM monoclonal antibody. Both antibodies recognized P0 expressed on the surface of peripheral nerve myelin and the P0 liposomes. P0 liposomes and peripheral nerve myelin treated with trypsin lost the surface determinant that reacted with the monoclonal antibody. Analysis of the trypsin-treated liposomes and peripheral nerve myelin by polyacrylamide gel electrophoresis revealed molecular weights for this protein of 19,500 and 20,500, respectively. Similar treatment of the P0 in the fluid phase resulted in many smaller fragments. These results indicate that P0 consists of two domains, a hydrophilic domain accessible to trypsin digestion and a hydrophobic domain, which is potentially trypsin-sensitive, but shielded by the lipid bilayer. Binding studies with an anti-P0 monoclonal antibody and polyacrylamide gel analysis of the lipid-shielded P0 fragment in liposomes and peripheral nerve myelin suggest that the orientation of the protein in the liposome is similar to that in peripheral nerve myelin.
D000906:Antibodies; D000936:Antigen-Antibody Complex; D002784:Cholesterol; D004134:Dimyristoylphosphatidylcholine; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007700:Kinetics; D008051:Lipid Bilayers; D008854:Microscopy, Electron; D018993:Myelin P0 Protein; D009185:Myelin Proteins; D009186:Myelin Sheath / Q000032:analysis; D012584:Sciatic Nerve / Q000032:analysis
D000906:Antibodies; D000936:Antigen-Antibody Complex; D008051:Lipid Bilayers; D018993:Myelin P0 Protein; D009185:Myelin Proteins; D002784:Cholesterol; D004134:Dimyristoylphosphatidylcholine