Differences in epitope recognition, isotype and titer of antisera to Plasmodium falciparum merozoite surface protein 4 raised by different modes of DNA or protein immunization

Plasmodium falciparum merozoite surface protein 4 (MSP4) is being developed as a component of a subunit vaccine against asexual stages of malaria. Thr ee DNA constructs were produced that induced expression of MSP4 either in t he cytoplasm of transfected cells or secreted from cells under the control of the human tissue plasminogen activator (TPA) signal or the native P. fal ciparum MSP4 signal. Only the construct containing the TPA signal induced d etectable antibodies in mice, although gene expression was demonstrated in all constructs and MSP4 was shown to be secreted using either signal by in vitro transient transfection of COS cells. Two recombinant MSP4 proteins th at encoded the same sequence as the plasmid DNA were produced in E. coli (E cMSP4-His) and S. cerevisiae (yMSP4-His) and used to raise antibodies in mi ce. Comparison of the antibodies elicited by these various antigen formulat ions showed differences in titer, isotype and epitope recognition. The tite r of antibodies induced by DNA:vaccination was lower than that induced by y MSP4-His, which in turn was lower than that induced by EcMSP3-His. The isot ype profiles of the antibodies were also different, the plasmid DNA induced predominantly IgG, responses whereas the two proteins induced predominantl y IgG(1) responses. The antibodies induced by DNA and yMSP4-His recognized predominantly the C-terminal epidermal growth factor (EGF)-like domain of t he protein, whereas EcMSP3-His induced antibodies recognizing all domains o f the protein equally. The antibodies induced by DNA vaccination were direc ted almost extensively to conformational epitopes so that reactivity with n ative MSP4 was abolished after disulfide bonds in the protein were disrupte d. Antibodies induced by recombinant proteins recognized linear epitopes as well and reactivity to native MSP4 was preserved after reduction and alkyl ation of parasite proteins. (C) 2000 Elsevier Science Ltd. All rights reser ved.