Title : Unusual N-glycosylation of a recombinant human
erythropoietin expressed in a human lymphoblastoid cell line does not alter its biological properties
Abstract :
- Erythropoietin ( Epo ) is a 166 amino acids protein containing three N-glycosylation sites ( Asn-24, Asn-38, and Asn-83 ) and 1 O- glycosylation site ( Ser-126 ) and involved in the regulation of the level of red blood cells
- Today, only one recombinant human Epo ( rHuEpo ), produced in CHO cell line, is extensively used in therapy to cure severe anemia
- The structure of the glycan chains of this rHuEpo slightly differ of those of the urinary human Epo ( uHuEpo ), considered as the natural Epo molecule
- In an attempt to produce a rHuEpo as close as possible to the uHuEpo , Epo gene was expressed in a human lymphoblastoid cell line, named RPMI 1788
- In order to fully characterize the Epo-RPMI, structural characterizations of the protein skeleton as well as glycan chains were undergone
- As expected, the amino acid sequence of the Epo-RPMI conformed to that of uHuEpo
- Surprisingly, the structure of some N-glycan chains , as mainly determined by ESI-MS, revealed some unusual characteristics
- Thus, 80% of N-glycans possess a bisecting GlcNAc residue, 25% bear a second fucose residue which is present, in a large part, in a sialyl Le(x) motif , and 13% contain more than three LacNAc repeats (up to five per molecule)
- Despite these unusual structural characteristics, the data concerning the in vitro and in vivo biological activities were not impaired when compared to Epo-CHO and uHuEpo