more than 10-fold inhibition. Core protein variants with mutations in all phosphorylation sites exhibited dominant-negative effects on RNA encapsidation by wild-type protein. The results suggest that the presence of phosphoserine at position 162 of HBV core protein is required for pregenomic-RNA encapsidation, whereas phosphoserine at position 170 optimizes the process and serine might be preferable in position 155. Examination of the pregenomic-RNA-encapsidating capacities of DHBV core protein variants, in which four phosphorylation sites were jointly mutated to alanine or aspartic acid, suggests that phosphorylation of DHBV core protein at these sites may optimize pregenomic-RNA encapsidation but that its impact is much less profound than in the case of HBV. The possible mechanisms by which RNA encapsidation may be modulated by core protein phosphorylation are discussed in the context of the observed differences between the two viruses.The DNA genome of hepadnaviruses is replicated through reverse transcription of an RNA intermediate, the pregenomic RNA (reviewed in reference 4). Replication begins with encapsidation of pregenomic RNA, a process which requires two viral proteins, core (capsid) protein and polymerase; however, the enzymatic activities of the polymerase are not essential. Sequential synthesis of a minus-strand DNA by reverse transcription and synthesis of the second DNA strand are carried out by the polymerase inside the assembled nucleocapsid.

We analyzed expression of a putative oligopeptide permease (Opp) of Borrelia burgdorferi. Unlike the opp operons of other bacteria for which there is a single substrate binding protein, B. burgdorferi codes for three substrate binding proteins (OppA-I to -III) in its opp operon and an additional two homologs on plasmids (OppA-IV and -V). Instead of a single promoter region regulating transcription of the entire operon, as seen in other bacterial opp operons, it appears that among oppA-I, -II, and -III, as well as oppA-IV and -V, each has a potential upstream promoter region. We tested the function of these putative promoter sequences by fusion to a promoterless ␤-galactosidase reporter gene in pCB182. Each of the promoter regions was found to be active. The level of activity in the reporter constructs closely paralleled the level of expression of each gene in in vitro-grown B. burgdorferi. Changes in carbon and nitrogen availability differentially affected individual promoters, but no changes in promoter activity were seen when Escherichia coli bacteria (with the promoter constructs) were grown in various concentrations of phosphate and leucine and changes in pH. Expression of specific oppA genes with B. burgdorferi varied significantly between its mouse and fed and unfed tick hosts. Differences in regulation of opp gene expression suggest a potential role in environmental response by the organism.

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The three flavivirus glycoproteins prM, E and NS1 are formed by post-translational cleavage and are glycosylated by the addition of N-linked glycans. NS1 may form homodimers, whereas E may form homodimers, homotrimers or heterodimers (prM-E). Modification of these processes by mutagenesis of the proteins has the potential to generate viruses that are restricted in growth and are possible vaccine candidates. Using an SV40-based expression system, we previously analysed dimerization and secretion of the NS1 protein of dengue virus type 2 (DEN-2) with mutations in the conserved Cys residues, or within hydrophilic or hydrophobic regions, or at glycosylation sites. In this study, mutations which reduce cleavage at the DEN-2 prM/E signalase cleavage site are described. On the