The effect of protein synthesis inhibitors on the glycosylation site occupancy of recombinant human prolactin.
The relationship between synthesis and N-linked glycosylation site occupancy of recombinant human prolactin produced from C127 cells was studied with the aid of a battery of protein synthesis inhibitors. Non-lethal concentrations of sodium fluoride, gougerotin, puromycin, anisomycin, and emetine did not alter site occupancy, but low concentrations (< 10 micrograms ml-1) of cycloheximide increased the fraction of secreted prolactin bearing oligosaccharide from 20% to 80% of the total. Cycloheximide is an inhibitor of the elongation step of protein synthesis. The observed increase in glycosylation site occupancy upon addition of cycloheximide is consistent with the current opinion that the initial glycosylation event occurs cotranslationally during a limited time period. Cycloheximide may extend this time period by reducing elongation rate. However, the absence of any effect from treatment with other inhibitors of elongation suggests that cycloheximide is unique in its behavior on this system.
Shelikoff
M
M
Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge 02139-4308.
Sinskey
A J
AJ
Stephanopoulos
G
G
D000581:Amidohydrolases; D000595:Amino Acid Sequence; D000818:Animals; D001216:Asparagine; D015153:Blotting, Western; D002240:Carbohydrate Sequence; D002460:Cell Line; D004591:Electrophoresis, Polyacrylamide Gel; D006031:Glycosylation / Q000187:drug effects; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000737:chemistry / Q000378:metabolism; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D011388:Prolactin / Q000096:biosynthesis* / Q000302:isolation & purification; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000302:isolation & purification; D014162:Transfection
D009844:Oligosaccharides; D011500:Protein Synthesis Inhibitors; D011994:Recombinant Proteins; D001216:Asparagine; D011388:Prolactin; D000581:Amidohydrolases; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase