title: Structure-function analysis of heterodimer formation, oligomerization, and receptor binding of the Staphylococcus aureus bi-component toxin LukGH.
abstract: The bi-component leukocidins of Staphylococcus aureus are important virulence factors that lyse human phagocytic cells and contribute to immune evasion. The γ-hemolysins (HlgAB and HlgCB) and Panton-Valentine leukocidin (PVL or LukSF) were shown to assemble from soluble subunits into membrane-bound oligomers on the surface of target cells, creating barrel-like pore structures that lead to cell lysis. LukGH is the most distantly related member of this toxin family, sharing only 30-40% amino acid sequence identity with the others. We observed that, unlike other leukocidin subunits, recombinant LukH and LukG had low solubility and were unable to bind to target cells, unless both components were present. Using biolayer interferometry and intrinsic tryptophan fluorescence we detected binding of LukH to LukG in solution with an affinity in the low nanomolar range and dynamic light scattering measurements confirmed formation of a heterodimer. We elucidated the structure of LukGH by x-ray crystallography at 2.8-Å resolution. This revealed an octameric structure that strongly resembles that reported for HlgAB, but with important structural differences. Structure guided mutagenesis studies demonstrated that three salt bridges, not found in other bi-component leukocidins, are essential for dimer formation in solution and receptor binding. We detected weak binding of LukH, but not LukG, to the cellular receptor CD11b by biolayer interferometry, suggesting that in common with other members of this toxin family, the S-component has the primary contact role with the receptor. These new insights provide the basis for novel strategies to counteract this powerful toxin and Staphylococcus aureus pathogenesis.
authors.affiliation: From Arsanis Biosciences, Vienna Biocenter Campus, Helmut-Qualtinger-Gasse 2, 1030 Vienna, Austria and.
authors.forename: Adriana
authors.initials: A
authors.lastname: Badarau
authors.affiliation: From Arsanis Biosciences, Vienna Biocenter Campus, Helmut-Qualtinger-Gasse 2, 1030 Vienna, Austria and.
authors.forename: Harald
authors.initials: H
authors.lastname: Rouha
authors.affiliation: From Arsanis Biosciences, Vienna Biocenter Campus, Helmut-Qualtinger-Gasse 2, 1030 Vienna, Austria and.
authors.forename: Stefan
authors.initials: S
authors.lastname: Malafa
authors.affiliation: SARomics Biostructures AB, Medicon Village, S-223 81 Lund, Sweden.
authors.forename: Derek T
authors.initials: DT
authors.lastname: Logan
authors.affiliation: SARomics Biostructures AB, Medicon Village, S-223 81 Lund, Sweden.
authors.forename: Maria
authors.initials: M
authors.lastname: Håkansson
authors.affiliation: From Arsanis Biosciences, Vienna Biocenter Campus, Helmut-Qualtinger-Gasse 2, 1030 Vienna, Austria and.
authors.forename: Lukas
authors.initials: L
authors.lastname: Stulik
authors.affiliation: From Arsanis Biosciences, Vienna Biocenter Campus, Helmut-Qualtinger-Gasse 2, 1030 Vienna, Austria and.
authors.forename: Ivana
authors.initials: I
authors.lastname: Dolezilkova
authors.affiliation: From Arsanis Biosciences, Vienna Biocenter Campus, Helmut-Qualtinger-Gasse 2, 1030 Vienna, Austria and.
authors.forename: Astrid
authors.initials: A
authors.lastname: Teubenbacher
authors.affiliation: From Arsanis Biosciences, Vienna Biocenter Campus, Helmut-Qualtinger-Gasse 2, 1030 Vienna, Austria and.
authors.forename: Karin
authors.initials: K
authors.lastname: Gross
authors.affiliation: From Arsanis Biosciences, Vienna Biocenter Campus, Helmut-Qualtinger-Gasse 2, 1030 Vienna, Austria and.
authors.forename: Barbara
authors.initials: B
authors.lastname: Maierhofer
authors.affiliation: From Arsanis Biosciences, Vienna Biocenter Campus, Helmut-Qualtinger-Gasse 2, 1030 Vienna, Austria and.
authors.forename: Susanne
authors.initials: S
authors.lastname: Weber
authors.affiliation: From Arsanis Biosciences, Vienna Biocenter Campus, Helmut-Qualtinger-Gasse 2, 1030 Vienna, Austria and.
authors.forename: Michaela
authors.initials: M
authors.lastname: Jägerhofer
authors.affiliation: From Arsanis Biosciences, Vienna Biocenter Campus, Helmut-Qualtinger-Gasse 2, 1030 Vienna, Austria and.
authors.forename: David
authors.initials: D
authors.lastname: Hoffman
authors.affiliation: From Arsanis Biosciences, Vienna Biocenter Campus, Helmut-Qualtinger-Gasse 2, 1030 Vienna, Austria and eszter.nagy@arsanis.com.
authors.forename: Eszter
authors.initials: E
authors.lastname: Nagy
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D001665:Binding Sites; D039481:CD11b Antigen / Q000737:chemistry* / Q000378:metabolism; D018360:Crystallography, X-Ray; D018922:HL-60 Cells; D006460:Hemolysin Proteins / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D007956:Leukocidins / Q000737:chemistry* / Q000378:metabolism; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding; D055503:Protein Multimerization; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013211:Staphylococcus aureus / Q000737:chemistry* / Q000378:metabolism; D013329:Structure-Activity Relationship
chemical_list: D001426:Bacterial Proteins; D039481:CD11b Antigen; D006460:Hemolysin Proteins; C509295:ITGAM protein, human; D007956:Leukocidins; D011994:Recombinant Proteins; C586430:leukocidin AB, Staphylococcus aureus
keywords: Bacterial Toxin; Crystal Structure; Crystallography; Microbial Pathogenesis; Mutagenesis; Staphylococcus aureus (S. aureus)