Characterization of a highly glycosylated biosynthetic intermediate of ovalbumin.
Ovalbumin extracted from oviduct slices incubated with [35S]methionine or [2-3]mannose contained two biosynthetic intermediates, OE and OF (Y. Kato, H. Iwase, and K. Hotta, 1984, J. Biochem. (Tokyo) 95, 455-463). In the present study, these intermediates are further characterized. The 3H dpm/35S dpm ratio of OF labeled with both [35S]methionine and [3H]mannose was twice that or greater than that of OE. The tritium-labeled OF migrated more slowly than OE on sodium dodecyl sulfate-gel electrophoresis and had a molecular weight exceeding that of OE by about 1500. These findings, along with the fact that [3H]OF had sugar chains similar to those of [3H]OE, suggest that OF may possibly have two sugar chains in one molecule. For confirmation of this, the glycosylation sites of OF were examined. Peptic and tryptic glycopeptides were prepared from [2-3H]mannose-labeled OF and the other ovalbumin subfractions--OA, OC, OD, and OE--and then analyzed by high-performance liquid chromatography. The peptic glycopeptides prepared from [3H]OF contained glycopeptides in addition to those derived from the other subfractions although tryptic glycopeptides obtained from [3H]OF were similar to those from the other subfractions. This shows that the above hypothesis is valid.
Kato
Y
Y
Iwase
H
H
Hotta
K
K
D000118:Acetylglucosaminidase / Q000502:physiology; D000818:Animals; D002645:Chickens; D002847:Chromatography, Agarose; D002851:Chromatography, High Pressure Liquid; D005260:Female; D006020:Glycopeptides / Q000032:analysis*; D006023:Glycoproteins / Q000096:biosynthesis; D066298:In Vitro Techniques; D008358:Mannose / Q000378:metabolism; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D008715:Methionine / Q000378:metabolism; D010047:Ovalbumin / Q000096:biosynthesis*; D010057:Oviducts / Q000378:metabolism; D013462:Sulfur Radioisotopes; D014316:Tritium
D006020:Glycopeptides; D006023:Glycoproteins; D013462:Sulfur Radioisotopes; D014316:Tritium; D010047:Ovalbumin; D008715:Methionine; D000118:Acetylglucosaminidase; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D008358:Mannose