Analysis of the cellular localization of Bdr paralogs in Borrelia burgdorferi, a causative agent of Lyme disease: Evidence for functional diversity

The bdr (Borrelia direct repeat) gene family of the genus Borrelia encodes a polymorphic group of proteins that carry a central repeat motif region co ntaining putative phosphorylation sites and a hydrophobic carboxyl-terminal domain. It has been postulated that the Bdr proteins may anchor to the inn er membrane via the C-terminal domain. In this study, we used cellular frac tionation methodologies, salt and detergent treatments, and immunoblot anal yses to assess the association of the Bdr proteins with the cellular infras tructure in both Borrelia burgdorferi (a Lyme disease spirochete) and B. tu ricatae (a relapsing fever spirochete). Triton X-114 extraction and partiti oning experiments demonstrated that most Bdr paralogs are associated with t he inner membrane-peptidoglycan complex. Analyses of cells treated with the highly chaotropic bile salt detergent deoxycholic acid demonstrated that s ome Bdr paralogs may also interact with the peptidoglycan, as evidenced by their tight association with the insoluble cellular matrix. In addition, im munoprecipitation (IP) experiments revealed an enhanced IP of all Bdr paral ogs when the cell lysates were boiled prior to addition of the precipitatin g antibody. Furthermore, some Bdr paralogs were accessible to antibody in t he IP experiments only in the boiled cell lysates. These observations sugge st that different Bdr paralogs may carry out different structural-functiona l roles. Demonstration of the inner membrane localization of the Bdr protei ns and of the differences in nature of the interaction of individual Bdr pa ralogs with the cell infrastructure is an important step toward defining th e functional role of this unique protein family in the genus Borrelia.