Two Crystal Structures of Pneumococcal Pilus Sortase C Provide Novel Insights into Catalysis and Substrate Specificity |
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Authors: | Fabrice Neiers Chaithanya Madhurantakam Stefan Fälker Clothilde Manzano Staffan Normark Adnane Achour |
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Institution: | 1 Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, SE-171 77 Solna, Sweden 2 Center for Infectious Medicine, F59, Department of Medicine Huddinge, Karolinska University Hospital Huddinge, Karolinska Institutet, SE-141 86 Stockholm, Sweden 3 Department of Bacteriology, Swedish Institute for Infectious Disease Control, SE-171 82 Solna, Sweden 4 Institut de Biologie Structurale Jean-Pierre Ebel, UMR 5075 (CEA, CNRS, UJF, PSB), 41 rue Jules Horowitz, F-38027 Grenoble, France |
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Abstract: | The respiratory tract pathogen Streptococcus pneumoniae is a primary cause of morbidity and mortality worldwide. Pili enhance initial adhesion as well as the capacity of pneumococci to cause pneumonia and bacteremia. Pilus-associated sortases (SrtB, SrtC, and SrtD) are involved in the biogenesis of pneumococcal pili, composed of repeating units of RrgB that create the stalk to which the RrgA adhesin and the preferential pilus tip subunit RrgC are covalently associated. Using single sortase-expressing strains, we demonstrate that both pilin-polymerizing sortases SrtB and SrtC can covalently link pili to the peptidoglycan cell wall, a property shared with the non-pilus-polymerizing enzyme SrtD and the housekeeping sortase SrtA. Comparative analysis of the crystal structures of S. pneumoniae SrtC and SrtB revealed structural differences explaining the incapacity of SrtC, but not of SrtB, to incorporate RrgC into the pilus. Accordingly, site-directed mutagenesis of Thr160 in SrtB to an arginine as in SrtC (Arg160) partially converted its substrate specificity into that of SrtC. Solving two crystal structures for SrtC suggests that an opening of a flexible lid and a concomitant cysteine rotation are important for catalysis and the activation of the catalytic cysteine of pilus-associated sortases. |
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Keywords: | sortase crystal structure substrate recognition catalysis pili |
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