ORIGINAL PAPER Solution structure of the major (Spy0128) and minor (Spy0125 and Spy0130) pili subunits from Streptococcus pyogenes Alexandra S. Solovyova Æ Jonathan A. Pointon Æ Paul R. Race Æ Wendy D. Smith Æ Michael A. Kehoe Æ Mark J. Banfield Received: 14 November 2008 / Revised: 17 February 2009 / Accepted: 23 February 2009 / Published online: 17 March 2009 Ó European Biophysical Societies’ Association 2009 Abstract Adhesion of the serotype M1 Streptococcus pyogenes strain SF370 to human tonsil explants and cultured keratinocytes requires extended polymeric surface structures called pili. In this important human pathogen, pili are assembled from three protein subunits: Spy0125, Spy0128 and Spy0130 through the action of sortase enzymes. For this study, the structural properties of these pili proteins have been investigated in solution. Spy0125 and Spy0128 display characteristics of globular, folded proteins. Circular dichroism suggests a largely b-sheet composition for Spy0128 and Spy0125; Spy0130 appears to contain little secondary structure. Each of the proteins adopts a monodisperse, monomeric state in solution as assessed by analytical ultracentrifugation. Further, small- angle X-ray scattering curves for Spy0125, Spy0128 and Spy0130 suggest each protein adopts an elongated shape, likely comprised of two domains, with similar maximal dimensions. Based on the scattering data, dummy atom models of each of the pili subunits have been reconstructed ab initio. This study provides the first insights into the structure of Streptococcus pyogenes minor pili subunits, and possible implications for protein function are discussed. Keywords Pili subunits  Circular dichroism  Analytical ultracentrifugation  Small-angle X-ray scattering  Dummy atom model  Structural disorder Introduction Streptococcus pyogenes (also called Group A Streptococcus, or GAS) is an important Gram-positive bacterial pathogen that causes a wide variety of diseases in humans. These range in severity from mild self-limiting infections of the pharynx or skin, to more serious and potentially life-threatening infections such as severe pharyngitis, tonsillitis, strepto- coccal toxic-shock syndrome and necrotising fasciitis. A first critical step in the pathogenesis of many bacterial infections is specific adhesion to a particular host surface. A number of GAS cell-surface molecules have been implicated as specific adhesins by in vitro studies employing cultured mammalian cell lines, such as Hep-2 and A549 cells [reviewed in (Courtney et al. 2002)]. However, the extent to which these long-established cell-lines represent natural human host epithelial surfaces has been questioned (Abbot et al. 2007). Recent studies, using intact human tonsil explants and primary human keratinocytes, revealed that bacterial cell- surface structures called pili (also known as fimbriae) are required for efficient adhesion of the serotype M1 GAS strain SF370 (henceforth ‘‘GAS’’) to these clinically relevant host tissues (Abbot et al. 2007). Similar results were obtained from assessing the binding of GAS pilus-negative mutants to Detroit-562 pharyngeal cells (Edwards et al. 2008). A. S. Solovyova, J. A. Pointon and P. R. Race have contributed equally to this work. AUC&HYDRO 2008—Contributions from 17th International Symposium on Analytical Ultracentrifugation and Hydrodynamics, Newcastle, UK, 11–12 September 2008. A. S. Solovyova (&)  J. A. Pointon  P. R. Race  W. D. Smith  M. A. Kehoe Faculty of Medical Sciences, Institute for Cell and Molecular Biosciences, Newcastle University, Framlington Place, Newcastle upon Tyne NE2 4HH, UK e-mail: alexandra.solovyova@newcastle.ac.uk Present Address: M. J. Banfield Department of Biological Chemistry, John Innes Centre, Norwich Research Park, Norwich NR4 7UH, UK 123 Eur Biophys J (2010) 39:469–480 DOI 10.1007/s00249-009-0432-2