Identification of a major B-cell epitope of the Plasmodium falciparum glutamate-rich protein (GLURP), targeted by human antibodies mediating parasite killing.

The antigenicity of the glutamate-rich protein (GLURP) of Plasmodium falciparum was comprehensively evaluated in epitope-mapping studies utilizing a phage display library, synthetic peptides and anti-GLURP IgG preparations previously shown to promote strong antibody-dependent cellular inhibition (ADCI) effects. We identified six major B-cell epitopes within the nonrepetitive region R0, corresponding to amino acid residues 173 to 187 (P1), 193 to 207 (P3), 216 to 229 (P4), 264 to 288 (P11), 343 to 357 (P10), and 407 to 434 (S3). Of these, four (P1, P3, P4, and S3) were frequently recognized by high-titered IgG antibodies in plasma samples from immune Liberian adults (prevalence: 29.1-45.0%). The three epitopes P1, P3, and P4 contained a common motif (seven out of nine positions are identical) and may thus constitute a family of structurally related epitopes. This leaves two distinct epitopes, one (P3) representing this new epitope family and S3 as targets for biologically active antibodies. Human IgG antibodies from single plasma samples were affinity-purified against these peptides. P3-specific IgG preparations were consistently more effective in ADCI than S3-specific IgG. Among the different GLURP epitopes, we therefore suggest that the P3 epitope is potentially the most important epitope in GLURP for the development of clinical immunity to malaria in man.

Activation control of pur gene expression in Lactococcus lactis: proposal for a consensus activator binding sequence based on deletion analysis and site-directed mutagenesis of purC and purD promoter regions.

A comparison of the purC and purD upstream regions from Lactococcus lactis revealed the presence of a conserved ACCGAACAAT decanucleotide sequence located precisely between -79 and -70 nucleotides upstream from the transcriptional start sites. Both promoters have well-defined -10 regions but lack sequences resembling -35 regions for sigma70 promoters. Fusion studies indicated the importance of the conserved sequence in purine-mediated regulation. Adjacent to the conserved sequence in purC is a second and similar region required for high-level expression of the gene. A consensus PurBox sequence (AWWWCCGAACWWT) could be proposed for the three regions. By site-directed mutagenesis we found that mutation of the central G in the PurBox sequence to C resulted in low levels of transcription and the loss of purine-mediated regulation at the purC and purD promoters. Deletion analysis also showed that the nucleotides before the central CCGAAC core in the PurBox sequence are important. All results support the idea that purC and purD transcription is regulated by a transcriptional activator binding to the PurBox sequence.