Biophysical characterization and structural insights of leptospiral complement regulator-acquiring protein A.

Many pathogens establish a successful infection by evading the host complement system, an essential arm of innate immunity. Pathogenic Leptospira is reported to escape complement-mediated killing by recruiting the host complement regulators by lipoproteins or outer surface proteins. One of the outer surface proteins, Leptospiral complement regulator-acquiring protein A (LcpA), is known to recruit complement regulators, C4b-binding protein (C4BP), and Factor H (FH) on the bacterial surface. Mapping of interacting domains from C4BP and FH with the LcpA has already been reported. However, the region or structural part of the LcpA mediating the interaction is not known yet. Here, we report cloning, expression, refolding and purification of recombinant LcpA from an inclusion body of E. coli heterologous expression system. We also demonstrate the biophysical characterization of recombinant LcpA and reveal its secondary structure contents. Moreover, the protein displays a moderate thermostability. The change of intrinsic fluorescence and CD spectra demonstrate a change in the secondary structure of protein due to binding with Zn ions. Molecular docking of LcpA with the complement regulators displays important interface residues from both the individual counterparts. Molecular dynamic simulation analysis demonstrates the stability of interactions between LcpA and C4BP. In our understanding, this is the first report on the large-scale purification of LcpA through refolding experiments and biophysical characterization of LcpA. This study may provide additional information on the structural basis of binding with the complement regulators.