Figure 2.

BB0238 is a subsurface protein that exists in the periplasm and as a membrane-bound form and is produced throughout the tick-mouse infection cycle of Borrelia burgdorferi. A, BB0238 is not exposed on the spirochete surface. B, burgdorferi cells were incubated with (+) or without (−) proteinase K for the removal of protease-sensitive surface proteins and then processed for immunoblot analysis using either BB0238 or control surface-exposed (OspA) and subsurface (FlaB) protein antibodies. B, BB0238 is undetectable in the isolated outer membrane vesicle fraction. B. burgdorferi lysates were separated into protoplasmic cylinder (PC) and outer membrane vesicle (OMV) fractions by sodium dodecyl sulfate–polyacrylamide gel electrophoresis and immunoblotted with antiserum against BB0238 or outer membrane protein (OspA). C, Detection of BB0238 in the periplasmic and spheroplasmic fractions of B. burgdorferi cells. Periplasmic or spheroplasmic proteins were extracted from cultured spirochetes and immunoblotted using BB0238 antibodies. D, Triton X-114 phase partitioning of B. burgdorferi proteins. Spirochete lysates were subjected to Triton X-114 phase partitioning of aqueous and detergent phases and immunoblotted using antibodies against BB0238, N-terminal BB0323, C-terminal BB0323, or a control (OspA) protein. E, Consistent expression of bb0238 during the enzootic life cycle of the spirochete. Total tissue RNA was isolated from representative stages of murine and tick infection, and bb0238 transcripts were measured using quantitative reverse-transcription polymerase chain reaction (qRT-PCR) and presented as copies of bb0238 transcript per copy of flaB transcript. Error bars show the means and standard errors of the mean from ≥3 independent qRT-PCR analyses from individual ticks.