Glycosylation and proteolytic processing of 70 kDa C-terminal recombinant polypeptides of Plasmodium falciparum merozoite surface protein 1 expressedin mammalian cells

The cDNAs that encode the 70 kDa C-terminal portion of Plasmodium falciparu m merozoite surface protein 1 (MSP-1)! with or without an N-terminal signal peptide sequence and C-terminal glycosglphosphatidylinositol (GPI) signal sequence of MSP-1, were expressed in mammalian cell lines via recombinant v accinia virus. The polypeptides were studied with respect to the nature of glycosylation, localization, and proteolytic processing. The polypeptides d erived from the cDNAs that contained the N-terminal signal peptide were mod ified with N-linked high mannose type structures and low levels of O-linked oligosaccharides, whereas the polypeptides from the cDNAs that lacked the signal peptide were not glycosylated, The GPI anchor moiety is either absen t or present at a very low level in the polypeptide expressed from the cDNA that contained both the signal peptide and GPI signal sequences. Together, these data establish that whereas the signal peptide of MSP-1 is functiona l, the GPI anchor signal is either nonfunctional or poorly functional in ma mmalian cells. The polypeptides expressed from the cDNAs that contained the signal peptide were proteolytically cleaved at their C-termini, whereas th e polypeptides expressed from the cDNAs that lacked the signal peptide were uncleaved, While the polypeptide expressed from the cDNA containing both t he signal peptide and GPI anchor signal was truncated by similar to 14 kDa at the C-terminus, the polypeptide derived from the cDNA with only the sign al peptide was processed to remove similar to 6 kDa, also from the C-termin us, Furthermore, the polypeptides derived from cDNAs that lacked the signal peptide were exclusively localized intracellularly, the polypeptides from cDNAs that contained the signal peptide were predominantly intracellular, w ith low levels on the cell surface; none of the polypeptides was secreted i nto the culture medium to a detectable level, These results suggest that N- glycosylation alone is not sufficient for the efficient extracellular trans port of the recombinant MSP-1 polypeptides through the secretory pathway in mammalian cells.