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Assigned NMR backbone resonances of the ligand-binding region domain of the pneumococcal serine-rich repeat protein (PsrP-BR) reveal a rigid monomer in solution.

ABSTRACT: The pneumococcal serine rich repeat protein (PsrP) is displayed on the surface of Streptococcus pneumoniae with a suggested role in colonization in the human upper respiratory tract. Full-length PsrP is a 4000 residue-long multi-domain protein comprising a positively charged functional binding region (BR) domain for interaction with keratin and extracellular DNA during pneumococcal adhesion and biofilm formation, respectively. The previously determined crystal structure of the BR domain revealed a flat compressed barrel comprising two sides with an extended ?-sheet on one side, and another ?-sheet that is distorted by loops and ?-turns on the other side. Crystallographic B-factors indicated a relatively high mobility of loop regions that were hypothesized to be important for binding. Furthermore, the crystal structure revealed an inter-molecular ?-sheet formed between edge strands of two symmetry-related molecules, which could promote bacterial aggregation during biofilm formation. Here we report the near complete 15N/13C/1H backbone resonance assignment of the BR domain of PsrP, revealing a secondary structure profile that is almost identical to the X-ray structure. Dynamic 15N-T1, T2 and NOE data suggest a monomeric and rigid structure of BR with disordered residues only at the N- and C-termini. The presented peak assignment will allow us to identify BR residues that are crucial for ligand binding.

SUBMITTER: Schulte T

PROVIDER: S-EPMC7462905 | biostudies-literature | 2020 Oct

REPOSITORIES: biostudies-literature

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Assigned NMR backbone resonances of the ligand-binding region domain of the pneumococcal serine-rich repeat protein (PsrP-BR) reveal a rigid monomer in solution.

Schulte Tim T Sala Benedetta Maria BM Nilvebrant Johan J Nygren Per-Åke PÅ Achour Adnane A Shernyukov Andrey A Agback Tatiana T Agback Peter P

Biomolecular NMR assignments 20200420 2

The pneumococcal serine rich repeat protein (PsrP) is displayed on the surface of Streptococcus pneumoniae with a suggested role in colonization in the human upper respiratory tract. Full-length PsrP is a 4000 residue-long multi-domain protein comprising a positively charged functional binding region (BR) domain for interaction with keratin and extracellular DNA during pneumococcal adhesion and biofilm formation, respectively. The previously determined crystal structure of the BR domain revealed ...[more]

PMID: 32314099

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Project description:Pneumococcal pneumonia remains a WHO high-priority disease despite multivalent conjugate vaccines administered in clinical practice worldwide. A protein-based, serotype-independent vaccine has long-promised comprehensive coverage of most clinical isolates of the pneumococcus. Along with numerous pneumococcal surface protein immunogens, the pneumococcal serine-rich repeat protein (PsrP) has been investigated as a potential vaccine target due to its surface exposure and functions toward bacterial virulence and lung infection. Three critical criteria for its vaccine potential - the clinical prevalence, serotype distribution, and sequence homology of PsrP - have yet to be well characterized. Here, we used genomes of 13,454 clinically isolated pneumococci from the Global Pneumococcal Sequencing project to investigate PsrP presence among isolates, distribution among serotypes, and interrogate its homology as a protein across species. These isolates represent all age groups, countries worldwide, and types of pneumococcal infection. We found PsrP present in at least 50% of all isolates across all determined serotypes and nontypeable (NT) clinical isolates. Using a combination of peptide matching and HMM profiles built on full-length and individual PsrP domains, we identified novel variants that expand PsrP diversity and prevalence. We also observed sequence variability in its basic region (BR) between isolates and serotypes. PsrP has a strong vaccine potential due to its breadth of coverage, especially in nonvaccine serotypes (NVTs) when exploiting its regions of conservation in vaccine design. IMPORTANCE An updated outlook on PsrP prevalence and serotype distribution sheds new light on the comprehensiveness of a PsrP-based protein vaccine. The protein is present in all vaccine serotypes and highly present in the next wave of potentially disease-causing serotypes not included in the current multivalent conjugate vaccines. Furthermore, PsrP is strongly correlated with clinical isolates harboring pneumococcal disease as opposed to pneumococcal carriage. PsrP is also highly present in strains and serotypes from Africa, where the need for a protein-based vaccine is the greatest, giving new reasoning to pursue PsrP as a protein vaccine.

Project description:Streptococcus pneumoniae has evolved versatile strategies to colonize the nasopharynx of humans. Colonization is facilitated by direct interactions with host cell receptors or via binding to components of the extracellular matrix. In addition, pneumococci hijack host-derived extracellular proteases such as the serine protease plasmin(ogen) for ECM and mucus degradation as well as colonization. S. pneumoniae expresses strain-dependent up to four serine proteases. In this study, we assessed the role of secreted or cell-bound serine proteases HtrA, PrtA, SFP, and CbpG, in adherence assays and in a mouse colonization model. We hypothesized that the redundancy of serine proteases compensates for the deficiency of a single enzyme. Therefore, double and triple mutants were generated in serotype 19F strain EF3030 and serotype 4 strain TIGR4. Strain EF3030 produces only three serine proteases and lacks the SFP encoding gene. In adherence studies using Detroit-562 epithelial cells, we demonstrated that both TIGR4Δcps and 19F mutants without serine proteases or expressing only CbpG, HtrA, or PrtA have a reduced ability to adhere to Detroit-562 cells. Consistent with these results, we show that the mutants of strain 19F, which preferentially colonizes mice, abrogate nasopharyngeal colonization in CD-1 mice after intranasal infection. The bacterial load in the nasopharynx was monitored for 14 days. Importantly, mutants showed significantly lower bacterial numbers in the nasopharynx two days after infection. Similarly, we detected a significantly reduced pneumococcal colonization on days 3, 7, and 14 post-inoculations. To assess the impact of pneumococcal serine proteases on acute infection, we infected mice intranasally with bioluminescent and invasive TIGR4 or isogenic triple mutants expressing only CbpG, HtrA, PrtA, or SFP. We imaged the acute lung infection in real-time and determined the survival of the mice. The TIGR4lux mutant expressing only PrtA showed a significant attenuation and was less virulent in the acute pneumonia model. In conclusion, our results showed that pneumococcal serine proteases contributed significantly to pneumococcal colonization but played only a minor role in pneumonia and invasive diseases. Because colonization is a prerequisite for invasive diseases and transmission, these enzymes could be promising candidates for the development of antimicrobials to reduce pneumococcal transmission.

Dataset Information

Assigned NMR backbone resonances of the ligand-binding region domain of the pneumococcal serine-rich repeat protein (PsrP-BR) reveal a rigid monomer in solution.

Publications

Assigned NMR backbone resonances of the ligand-binding region domain of the pneumococcal serine-rich repeat protein (PsrP-BR) reveal a rigid monomer in solution.

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