Expression of the @entity375 glycoprotein invertase in XXXX fibroblasts: glycosylation, secretion, and enzymatic activity.

@entity19 @entity2425 @entity73145 @entity1735 @entity498 @entity375 @entity12215
Oligosaccharide processing is controlled by host- and protein-dependent factors. To increase our understanding of the relative contribution of those factors we studied the glycosylation of @entity375 invertase expressed in a heterologous system. Invertase synthesized in @entity19 cells (an NIH 3T3-derived packaging line) is secreted efficiently, enzymatically active, and heavily glycosylated. It was estimated that the protein contains 8 or 9 @entity1735 chains. Two classes can be observed, of an approximate size of 100-110 kDa and 115-130 kDa, respectively. The size differences are due to differences in glycosylation. The smaller class contains two high-mannose @entity1735 chains; the remainder is of the complex type, sialylated and most likely tri- or tetraantennary. This profile parallels the situation observed with invertase glycosylation in @entity375 , where 2 of 9 or 10 chains remain unprocessed. The larger size class of invertase expressed in @entity19 fibroblasts has a different profile, since it contains probably only complex-type glycans. There are no apparent differences, however, in the size of the protein backbone between the two size classes. When invertase is synthesized in the presence of the mannosidase inhibitor @entity73145 , processing is blocked completely, since all glycans are susceptible to @entity498 . The glucosidase inhibitor @entity12215 does not inhibit processing completely. In both cases secretion of the protein is not affected. The glycosylation inhibitor @entity2425 prevents secretion of invertase completely when cells are cultured at 37 degrees C. At 26 degrees C, however, nonglycosylated invertase can be detected in the medium. These data suggest that glycosylation of invertase seems to be essential for the early steps of the secretory pathway but is less critical for later events.
