Molecular and evolutionary analysis of Borrelia burgdorferi 297 circular plasmid-encoded lipoproteins with OspE- and OspF-like leader peptides

Abstract

We previously described two OspE and three OspF homologs in Borrelia burgdorferi 297 (D. R. Akins, S. F. Porcella, T. G. Popova, D. Shevchenko, S. I. Baker, M. Li, M. V. Norgard, and J. D. Radolf, Mol. Microbiol. 18:507-520, 1995; D. R. Akins, K. W. Bourell, M. J. Caimano, M. V. Norgard, and J. D. Radolf, J. Clin. Investig. 101:2240-2250, 1998). In this study, we characterized four additional lipoproteins with OspE/F-like leader peptides (Elps) and demonstrated that all are encoded on plasmids homologous to cp32 and cp18 from the B31 and N40 strains, respectively. Statistical analysis of sequence similarities using the binary comparison algorithm revealed that the nine lipoproteins from strain 297, as well as the OspE, OspF, and Erp proteins from the N40 and B31 strains, fall into three distinct families. Based upon the observation that these lipoproteins all contain highly conserved leader peptides, we now propose that the ancestors of each of the three families arose from gene fusion events which joined a common N terminus to unrelated proteins. Additionally, further sequence analysis of the strain 297 circular plasmids revealed that rearrangements appear to have played an important role in generating sequence diversity among the members of these three families and that recombinational events in the downstream flanking regions appear to have occurred independently of those within the lipoprotein-encoding genes. The association of hypervariable regions with genes which are differentially expressed and/or subject to immunological pressures suggests that the Lyme disease spirochete has exploited recombinatorial processes to foster its parasitic strategy and enhance its immunoevasiveness.

FIG. 1

Schematic representation of the seven circular plasmid loci characterized in B. burgdorferi 297 containing ospE and ospF homologs and elp genes. ORFs are shown as boxed regions. The numbers above the ORFs in the downstream regions flanking the lipoprotein-encoding genes indicate the deduced sizes of the encoded polypeptides. The arrows below the ORFs indicate the directions of transcription. H indicates the unique HindIII sites for each locus. The dashed regions indicate unsequenced regions. The OspE, p21, OspF, BbK2.10, BbK2.11, and Elp proteins determined to be evolutionarily related are shaded the same. ORFs identified downstream of the lipoprotein-encoding loci which are highly similar between the different circular plasmids also are shaded the same.

FIG. 2

Dendrogram analysis of B. burgdorferi 297, B31, and N40 proteins with OspE/F-like leader peptides. Dendrogram analysis of the B. burgdorferi 297, B31, and N40 lipoproteins identified by BLAST searches using the B. burgdorferi 297 OspE/F homologs and Elp proteins. The dendrogram was generated by using the PILEUP program from the University of Wisconsin GCG sequence analysis software package, version 8.1. The B. burgdorferi B31 ErpI and ErpJ lipoproteins and the strain ZS7 pG lipoprotein were not included in the analysis because they are equivalent to ErpA, ErpB2, and ErpG, respectively (8, 39).

FIG. 3

Evidence for possible genetic rearrangements revealed by sequence alignments of the B. burgdorferi 297 OspE and OspF homologs and Elp proteins. ClustalW alignments of OspE and p21 (A); OspF, BbK2.10, and BbK2.11 (B); and ElpA1, ElpA2, ElpB1, and ElpB2 (C). In panel A, arrows and boldface type indicate tandem duplication and possible insertion-deletion, respectively. In panel C, boldface sequences indicate potential insertions-deletions described in text, while tandem repeats in ElpB2 are underlined. In all of the panels, identical and similar residues are shaded.