Structural studies of Campylobacter jejuni: Virulence proteins (Cj0977 and JlpA) and novel lipoproteins
Paek, Seonghee 1979-
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Campylobacter jejuni is a major cause of human gastroenteritis worldwide; however, its pathogenesis is not understood well. Cj0977, a cytoplasmic protein regulated by the flagellar promoter s28, and JlpA, a cell-surface exposed lipoprotein adhesin, are virulence factors, but their contributions in pathogenesis of C. jejuni remains elusive. Since there is no clue for these two protein structures, we aimed to discover structure-function relationship of these proteins. My work focused on establishing expression system, purification method, and crystallization method of these proteins, leading up to obtaining high quality crystals for successful X-ray diffraction. Several techniques were employed to yield diffracting crystals: limited proteolysis and stability test for Cj0977 and introduction of two additional Met sites within the JlpA sequence. Here we present the first view of the crystal structures of Cj0977 and JlpA. Cj0977 adopts a ‘hot dog’ fold, a famous protein folds found in numerous CoA derivative binding enzymes. The Cj0977 structure suggests its possible function as an acyl-CoA binding regulatory protein. The JlpA structure reveals a novel fold of unclosed half β-barrel and is reminiscent of other bacterial lipoproteins. The JlpA structure suggests that JlpA may accommodate multiple ligands and that a similar role for JlpA as a carrier of as yet unidentified Campylobacter-specific lipids. The unique fold of JlpA led us to initiate other C. jejuni lipoprotein study. Most of C. jejuni lipoproteins are unknown due to no apparent sequence homology in the sequence databank; however, they are considered to contribute to the pathogenesis. We aimed to uncover the crystal structures of C. jejuni lipoproteins to provide framework for investigating their functions. My work focused on four lipoproteins: Cj0090, Cj1026c, Cj1090c, and Cj1649. Among these, the Cj0090 structure reveals a novel variant of the immunoglobulin fold with β-sandwich architecture, suggesting that Cj0090 may be involved in protein-protein interactions, consistent with a possible role for bacterial lipoproteins. In addition, a phage display technique was performed to identify ligands for our target lipoproteins. The crystal structures revealed in this study contribute to provide clues for possible roles of our target proteins and enable to perform further structure-based functional analyses.