Iron is an essential nutrient for almost all living organisms andexerts central functions in the metabolism thanks to its favorablechemical properties. During infection,Staphylococcus aureusrelies on host iron reservoir exploiting hemoglobin (Hb) heme asthe preferred primary source. Hemophores deputed to hemeretrieval and internalization are expressed on bacterial surfaceand IsdB performs the first step, intercepting free Hb andextracting heme. IsdB is a proven virulence factor highlyexpressed during staphylococcal infections, raising interest forthe potential development of new antimicrobials. We exploited cryo-EM single particle analysis, Small (SAXS) and Wide (WAXS) Angle X-ray Scattering and time-resolved (TR) mea-surements to gain a deeper insight into IsdB:Hb complex forma-tion and IsdB-mediated heme extraction mechanisms. To thisaim, we exploited three Hb ligation states to isolate key steps inheme extraction: carboxyHb and oxygenated Hb, resistant toheme removal, and oxidized Hb, the physiological IsdB sub-strate. Two structures representing the pre- and after extractionIsdB:Hb complexes were solved at 2.9 A and 5.8 A resolution: data indicates a specific initial binding of IsdB to Hbb-subunits,followed by Hb tetramer dimerization and binding of a secondIsdB molecule ona-chains (Previously published in De Bei Oet al. (2022) PNAS in press). SAXS and WAXS static measure-ments confirmed the stoichiometry of the complexes, while TR-WAXS allowed the identification of the structural dynamicevents occurring upon IsdB:Hb interaction and the definition ofa sequential model describing the observed kinetics. Also, TR-spectroscopy revealed that heme extraction starts only upon IsdBbinding on both Hb subunits. These results broadly improve ourcurrent understanding of IsdB structural and functional dynamics, thus supporting future studies aimed to impairS. aureusironacquisition, a promising target for the development of a newclass of antimicrobials.
Understanding iron hijacking by Staphylococcus aureus: a structural and mechanistic insight into the interaction of IsdB hemophore with human hemoglobin / De Bei, O; Marchetti, M; Ronda, L; Gianquinto, E; Levantino, M; Lazzarato, L; Chirgadze, Dy; Hardwick, Sw; Cooper, Lr; Cozzi, M; Faggiano, S; Spyrakis, F; Luisi, Bf; Campanini, B; Bettati, S. - In: FEBS OPENBIO. - ISSN 2211-5463. - 12:S1(2022), pp. P-02.1-010.159-P-02.1-010.159. (Intervento presentato al convegno The Biochemistry Global Summit, 25th IUBMB Congress, 46th FEBS Congress, 15th PABMB Congress tenutosi a Lisbon nel July 9–14, 2022) [10.1002/2211-5463.13440].
Understanding iron hijacking by Staphylococcus aureus: a structural and mechanistic insight into the interaction of IsdB hemophore with human hemoglobin
De Bei, O;Marchetti, M;Ronda, L;Cozzi, M;Faggiano, S;Spyrakis, F;Luisi, BF;Campanini, B;Bettati, S
2022-01-01
Abstract
Iron is an essential nutrient for almost all living organisms andexerts central functions in the metabolism thanks to its favorablechemical properties. During infection,Staphylococcus aureusrelies on host iron reservoir exploiting hemoglobin (Hb) heme asthe preferred primary source. Hemophores deputed to hemeretrieval and internalization are expressed on bacterial surfaceand IsdB performs the first step, intercepting free Hb andextracting heme. IsdB is a proven virulence factor highlyexpressed during staphylococcal infections, raising interest forthe potential development of new antimicrobials. We exploited cryo-EM single particle analysis, Small (SAXS) and Wide (WAXS) Angle X-ray Scattering and time-resolved (TR) mea-surements to gain a deeper insight into IsdB:Hb complex forma-tion and IsdB-mediated heme extraction mechanisms. To thisaim, we exploited three Hb ligation states to isolate key steps inheme extraction: carboxyHb and oxygenated Hb, resistant toheme removal, and oxidized Hb, the physiological IsdB sub-strate. Two structures representing the pre- and after extractionIsdB:Hb complexes were solved at 2.9 A and 5.8 A resolution: data indicates a specific initial binding of IsdB to Hbb-subunits,followed by Hb tetramer dimerization and binding of a secondIsdB molecule ona-chains (Previously published in De Bei Oet al. (2022) PNAS in press). SAXS and WAXS static measure-ments confirmed the stoichiometry of the complexes, while TR-WAXS allowed the identification of the structural dynamicevents occurring upon IsdB:Hb interaction and the definition ofa sequential model describing the observed kinetics. Also, TR-spectroscopy revealed that heme extraction starts only upon IsdBbinding on both Hb subunits. These results broadly improve ourcurrent understanding of IsdB structural and functional dynamics, thus supporting future studies aimed to impairS. aureusironacquisition, a promising target for the development of a newclass of antimicrobials.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
