title: Characterizing the conformational dynamics of metal-free PsaA using molecular dynamics simulations and electron paramagnetic resonance spectroscopy.
abstract: Prokaryotic metal-ion receptor proteins, or solute-binding proteins, facilitate the acquisition of metal ions from the extracellular environment. Pneumococcal surface antigen A (PsaA) is the primary Mn(2+)-recruiting protein of the human pathogen Streptococcus pneumoniae and is essential for its in vivo colonization and virulence. The recently reported high-resolution structures of metal-free and metal-bound PsaA have provided the first insights into the mechanism of PsaA-facilitated metal binding. However, the conformational dynamics of metal-free PsaA in solution remain unknown. Here, we use continuous wave electron paramagnetic resonance (EPR) spectroscopy and molecular dynamics (MD) simulations to study the relative flexibility of the structural domains in metal-free PsaA and its distribution of conformations in solution. The results show that the crystal structure of metal-free PsaA is a good representation of the dominant conformation in solution, but the protein also samples structurally distinct conformations that are not captured by the crystal structure. Further, these results suggest that the metal binding site is both larger and more solvent exposed than indicated by the metal-free crystal structure. Collectively, this study provides atomic-resolution insight into the conformational dynamics of PsaA prior to metal binding and lays the groundwork for future EPR and MD based studies of PsaA in solution.
authors.affiliation: School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, Australia; Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia.
authors.forename: Evelyne
authors.initials: E
authors.lastname: Deplazes
authors.affiliation: Research Centre for Infectious Diseases, School of Biological Sciences, University of Adelaide, Adelaide, Australia.
authors.forename: Stephanie L
authors.initials: SL
authors.lastname: Begg
authors.affiliation: Henry Wellcome Unit for Biological EPR, School of Chemistry, Norwich Research Park, University of East Anglia, Norwich, UK.
authors.forename: Jessica H
authors.initials: JH
authors.lastname: van Wonderen
authors.affiliation: Research Centre for Infectious Diseases, School of Biological Sciences, University of Adelaide, Adelaide, Australia.
authors.forename: Rebecca
authors.initials: R
authors.lastname: Campbell
authors.affiliation: School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, Australia; Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia; Australian Infectious Diseases Research Centre, University of Queensland, Brisbane, Australia.
authors.forename: Bostjan
authors.initials: B
authors.lastname: Kobe
authors.affiliation: Research Centre for Infectious Diseases, School of Biological Sciences, University of Adelaide, Adelaide, Australia.
authors.forename: James C
authors.initials: JC
authors.lastname: Paton
authors.affiliation: Henry Wellcome Unit for Biological EPR, School of Chemistry, Norwich Research Park, University of East Anglia, Norwich, UK.
authors.forename: Fraser
authors.initials: F
authors.lastname: MacMillan
authors.affiliation: Research Centre for Infectious Diseases, School of Biological Sciences, University of Adelaide, Adelaide, Australia. Electronic address: christopher.mcdevitt@adelaide.edu.au.
authors.forename: Christopher A
authors.initials: CA
authors.lastname: McDevitt
authors.affiliation: School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, Australia; Research School of Chemistry, The Australian National University, Canberra, Australia. Electronic address: megan.o'mara@anu.edu.au.
authors.forename: Megan L
authors.initials: ML
authors.lastname: O'Mara
mesh_terms: D018829:Adhesins, Bacterial / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D004578:Electron Spin Resonance Spectroscopy; D008074:Lipoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008345:Manganese / Q000737:chemistry; D008670:Metals / Q000737:chemistry; D056004:Molecular Dynamics Simulation; D016296:Mutagenesis; D017434:Protein Structure, Tertiary; D013113:Spin Labels
chemical_list: D018829:Adhesins, Bacterial; D008074:Lipoproteins; D008670:Metals; C085000:PsaA protein, Streptococcus; D013113:Spin Labels; D008345:Manganese
keywords: Cluster A–I solute-binding protein (SBP); Electron paramagnetic resonance (EPR) spectroscopy; Molecular dynamics simulations; Protein flexibility; PsaA; Spin-label mobility; Streptococcus pneumoniae