Kinetics of Entry of P0 Protein into Peripheral Nerve Myelin

Sciatic nerves from 9-day-old rat pups were removed, sliced into 0.4-mm sections, and incubated with [3H]fucose or [14C]glycine precursors. The nerve slices system gave nearly linear incorporation of [3H]fucose as a function of time for 3 h, after an initial lag of approximately 30 min for homogenate and approximately 60 min for myelin. Incorporation of [3H]fucose at constant specific radioactivity was directly proportional to exogenous fucose levels over the range 3.0 x 10(-8) M to 1.5 x 10(-6) M. Analysis of labeled proteins by sodium dodecyl sulfate polyacrylamide gel electrophoresis showed that greater than 50% of labeled glycoprotein was P0, with no other major constituents. This system was used in fucose-chase experiments to determine that a period of approximately 20 min elapses between fucosylation and assembly of P0 into myelin. Cycloheximide inhibition of protein synthesis was used to determine that a period of approximately 33 min elapses between protein synthesis and appearance of P0 myelin.