Legionella pneumophilamacrophage infectivity potentiator protein appendage domains modulate protein dynamics and inhibitor binding

Abstract

Macrophage infectivity potentiator (MIP) proteins are widespread in human pathogens includingLegionella pneumophila, the causative agent of Legionnaires disease and protozoans such asTrypanosoma cruzi. All MIP proteins contain a FKBP (FK506 binding protein)-like prolyl-cis/trans-isomerase domain that hence presents an attractive drug target. Some MIPs such as the Legionella protein (LpMIP) have additional appendage domains of mostly unknown function. In full-length, homodimeric LpMIP, the N-terminal dimerization domain is linked to the FKBP-like domain via a long, free-standing stalk helix. Combining X-ray crystallography, NMR and EPR spectroscopy and SAXS, we elucidated the importance of the stalk helix for protein dynamics and inhibitor binding to the FKBP-like domain and bidirectional crosstalk between the different protein regions. The first comparison of a microbial MIP and a human FKBP in complex with the same synthetic inhibitor was made possible by high-resolution structures of LpMIP with a [4.3.1]-aza-bicyclic sulfonamide and provides a basis for designing pathogen-selective inhibitors. Through stereospecific methylation, the affinity of inhibitors to toL. pneumophilaandT. cruziMIP was greatly improved. The resulting X-ray inhibitor-complex structures of LpMIP and TcMIP at 1.49 and 1.34 Å, respectively, provide a starting point for developing potent inhibitors against MIPs from multiple pathogenic microorganisms.

[4.3.1]Bicyclic FKBP Ligands Inhibit Legionella Pneumophila Infection by LpMip‐Dependent and LpMip‐Independent Mechanisms**

“…[21] Furthermore, our structural data recently confirmed the conserved binding mode between LpMip and human FKBPs for [4.3.1]-bicyclic sulfonamides. [22] Building on this discovery, we screened our whole library consisting of over 1000 FKBP-focused compounds, featuring five major scaffolds for affinity towards LpMip, including the before mentioned [4.3.1]-bicyclic sulfonamide scaffold to determine the favored scaffold. We further investigated selected compounds in a set of biochemical and infection assays to refine our understanding of the mechanisms underlying the anti-virulence effects.…”