The anchoring of the surface proteins to the cell wall in gram-positive bacteria involves a peptide ligation reaction catalyzed by transpeptidase sortase. Most bacterial genomes encode multiple sortases with dedicated functions. Streptococcus pneumoniae (Sp) carries four sortases; a housekeeping sortase (SrtA), and three pilin specific sortases (SrtC1, C2, C3) dedicated to the biosynthesis of covalent pilus. Interestingly, SrtA, meant for performing housekeeping roles, is also implicated in pilus assembly of Sp. The allegiance of SpSrtA to the pathogenic pilus assembly makes it an ideal target for clinical inhibitor development. In this paper, we describe biochemical characterization, crystal structure and peptide substrate preference of SpSrtA. Transpeptidation reaction with a variety of substrates revealed that the enzyme preferred elongated LPXTG sequences and transferred them equally well to both Ala- and Gly-terminated peptides. Curiously, the crystal structure of both wild type and an active site (Cys to Ala) mutant of SpSrtA displayed inter-twined 3D-swapped dimers in which each protomer generated a classic eight-stranded beta-barrel ‘sortase fold'. Size-exclusion chromatography and sedimentation equilibrium measurements revealed the predominant presence of a dimer in equilibrium with its monomer. The crystal structure-based Cys–Cys distance mapping with defined chemical cross-linkers established the existence of 3D-swapped structure in solution. The swapping in SpSrtA, unprecedented for sortase family, may be physiologically relevant and meant to perform regulatory functions.
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December 2020
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The cover image shows the ternary complex structure of pol? incorporating dCTP* opposite templating xanthine. In this issue Jung and colleagues (pp. 4797–4810) demonstrate the solved structure of human DNA polymerase η (ploη) catalyzing across xanthine and hypoxanthine. Image courtesy of Seongmin Lee.
Research Article|
December 18 2020
Interrogation of 3D-swapped structure and functional attributes of quintessential Sortase A from Streptococcus pneumoniae
Tora Biswas;
Tora Biswas
*
Investigation, Methodology, Writing - original draft, Writing - review & editing
1National Institute of Immunology, Delhi 110067, India
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Anurag Misra;
Anurag Misra
*
2Department of Physics, Indian Institute of Science, Bangalore 560012, India
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Sreetama Das;
Sreetama Das
*
Investigation, Methodology, Writing - original draft, Writing - review & editing
2Department of Physics, Indian Institute of Science, Bangalore 560012, India
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Prity Yadav;
Prity Yadav
Investigation, Methodology, Writing - review & editing
1National Institute of Immunology, Delhi 110067, India
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Suryanarayanarao Ramakumar;
Conceptualization, Resources, Supervision, Funding acquisition, Project administration, Writing - review & editing
2Department of Physics, Indian Institute of Science, Bangalore 560012, India
Correspondence: Rajendra P. Roy (rproy@nii.ac.in) or Suryanarayanarao Ramakumar (ramak@iisc.ac.in)
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Rajendra P. Roy
1National Institute of Immunology, Delhi 110067, India
Correspondence: Rajendra P. Roy (rproy@nii.ac.in) or Suryanarayanarao Ramakumar (ramak@iisc.ac.in)
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Biochem J (2020) 477 (24): 4711–4728.
Article history
Received:
August 13 2020
Revision Received:
November 18 2020
Accepted:
November 24 2020
Accepted Manuscript online:
November 25 2020
Citation
Tora Biswas, Anurag Misra, Sreetama Das, Prity Yadav, Suryanarayanarao Ramakumar, Rajendra P. Roy; Interrogation of 3D-swapped structure and functional attributes of quintessential Sortase A from Streptococcus pneumoniae. Biochem J 23 December 2020; 477 (24): 4711–4728. doi: https://doi.org/10.1042/BCJ20200631
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