BIOLOGICAL AND PHYSICOCHEMICAL CHARACTERIZATION OF RECOMBINANT HUMAN ERYTHROPOIETINS FRACTIONATED BY MONO-Q COLUMN CHROMATOGRAPHY AND THEIRMODIFICATION WITH SIALYLTRANSFERASE

Ten erythropoietin (EPO) fractions differing in sialic acid content, r anging from 9.5 to 13.8 mol mol(-1) of EPO, were obtained from baby ha mster kidney cell-derived recombinant human EPO by Mono Q column chrom atography. The mean pI values of the EPO fractions determined by IEF-g el electrophoresis systematically shifted from 4.11 to 3.31, coincidin g with the sialic acid content, without a change in the constitution o f asialo N-linked oligosaccharides of each fraction. Although a linear relationship between the in vivo bioactivity and the sialic acid cont ent of the fractionated samples was observed until 12.1 mol mol(-1) of EPO, there was no further increase in their activity over 12.4 mol mo l(-1) of EPO. On the other hand, an inverse relationship between the i n vitro bioactivity and sialic acid content of EPO was observed. Also, we showed that the in vivo bioactivity of some fractions with low sia lic acid contents was increased after treatment with alpha 2,6-sialylt ransferase, but the in vivo bioactivity of the other fractions with hi gh sialic acid contents was either decreased or not affected.