Analysis of Borrelia burgdorferi Proteome and Protein–Protein Interactions
The proteome of Borrelia burgdorferi undergoes dynamic alterations as the microbe cycles through and persists in diverse host or vector environments. Therefore, studies of B. burgdorferi proteome and protein–protein interactions, which play central roles in biological processes in diverse organisms, are critical in understanding biology and infectivity of spirochetes. Here, we describe the proteomic analysis of B. burgdorferi by two-dimensional (2-D) gel electrophoresis followed by protein identification via liquid chromatography-mass spectrometry and database searching. We also describe assays for studying the interaction between borrelial proteins: a novel high-throughput luciferase assay, yeast two-hybrid assay, and a far-Western assay that are routinely used in our laboratories.
Key wordsProteomics analysis Two-dimensional (2-D) gel electrophoresis Mass spectrometry Protein–protein interaction Borrelia burgdorferi Luciferase assay Yeast two-hybrid Far-Western
We sincerely thank our collaborators and past and present members of our laboratory, especially Manish Kumar, Adam Coleman, Toru Kariu, Quentin Bernard, Brian Backstedt, and Kavita Sharma for their assistance with developing the protocols presented in this chapter. This work was supported by funding from University of Maryland, College Park as well as grants from the National Institute of Allergy and Infectious Diseases, Award Numbers AI080615, AI106059, and AI116620 to UP.
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