In vivo incorporation of [3H]palmitic acid into PO protein, the major intrinsic protein of rat sciatic nerve myelin.
Separation of rat sciatic nerve myelin proteins by sodium dodecyl sulfate-slab gel electrophoresis 18 h after injection of [3H]palmitic acid into the nerve demonstrated acylation of the PO protein. When sciatic nerve myelin labeled with [3H]palmitic acid was extracted with acidified chloroform/methanol or chloroform/methanol, the radioactivity associated with PO was retained. These results provided evidence that the radioactivity derived from [3H]palmitic acid in PO protein was not due to labeled phospholipid bound to PO protein by strong physical interaction. Furthermore, the radioactivity associated with purified preparations of PO remained after dialysis and re-electrophoresis, providing additional evidence that [3H]palmitic acid was firmly bound to PO. Treatment of myelin proteins with hydroxylamine at pH 6.6 released most of the radioactivity, indicating that [3H]palmitic acid was covalently bound by ester linkage to PO. Cleavage of purified acylated PO with methanolic NaOH released 85% of the protein-bound radioactivity. Gas-liquid chromatography of the fatty acids released from PO showed labeling of methyl esters of palmitate, stearate, and oleate (56, 16, and 5%, respectively). In addition, a small unlabeled peak with retention time identical with methyl linoleate was also observed. Intraneural injection of a mixture of [3H]palmitic acid and [14C]fucose or [3H]palmitic acid and [35S]sulfate in vivo followed by disc gel electrophoresis and determination of incorporated radioactivity clearly showed that PO was glycosylated, sulfated, and acylated. The potential significance of fatty acids linked to PO is discussed.
D000215:Acylation; D000818:Animals; D005643:Fucose / Q000378:metabolism; D018993:Myelin P0 Protein; D009185:Myelin Proteins / Q000302:isolation & purification / Q000378:metabolism*; D009186:Myelin Sheath / Q000032:analysis; D019308:Palmitic Acid; D010169:Palmitic Acids / Q000032:analysis*; D051381:Rats; D011919:Rats, Inbred Strains; D012584:Sciatic Nerve / Q000378:metabolism*; D013431:Sulfates / Q000378:metabolism; D014316:Tritium
D018993:Myelin P0 Protein; D009185:Myelin Proteins; D010169:Palmitic Acids; D013431:Sulfates; D014316:Tritium; D005643:Fucose; D019308:Palmitic Acid