Dataset Information

Ferritin Degradation by Pneumococcal HtrA, RadA and ClpP Serine Proteases : A Probable Way For Releasing and Acquisition Of Iron.

ABSTRACT: Purpose:Iron is a necessary element for the growth of bacteria; however, there are limited iron sources known for these microorganisms yet. Intracellular iron is stored as ferritin from, which releases iron in a gradual and controlled manner. The present study aimed to characterize ferritin-binding proteins (FBPs) of Streptococcus pneumoniae. Material and Methods:S. pneumoniae species were cultured in BHI broth containing ferritin (1094 ng/mL) for 4h at 37°C. Ferritin level was measured using ELISA assay. Bacterial proteome was electrophoresed on SDS-PAGE and then transferred on PVDF nitrocellulose membrane. Afterward, the PVDF membranes were incubated with a ferritin solution. Identification of ferritin binding proteins was performed using anti-ferritin monoclonal antibody conjugated with HRP enzyme. Molecular docking was used to assess the interaction between pneumococcal proteases and FBPs applying phenylmethylsulfonyl fluoride (PMSF) as a protease inhibitor. Results:No FBPs were identified in S. pneumoniae proteome. Moreover, ferritin levels have significantly (p<0.05) decreased following the growth of S. pneumoniae in ferritin-rich BHI medium. Also, molecular docking showed that RadA protease, ClpP hydrolase, and HtrA protease can potentially interact with PMSF protease inhibitors. On the other hand, the addition of the PMSF to the culture of S. pneumoniae prevented the reduction of ferritin, which indicates a potential role of RadA, ClpP, and HtrA proteases in ferritin degradation. Conclusion:Our results suggest that S. pneumoniae produces no FBPs and also cannot directly use ferritin as an iron source. However, ferritin may be degraded through a protease-mediated mode.

SUBMITTER: Kheirandish M

PROVIDER: S-EPMC7494395 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Ferritin Degradation by Pneumococcal HtrA, RadA and ClpP Serine Proteases : A Probable Way For Releasing and Acquisition Of Iron.

Kheirandish Mozhgan M Motlagh Behrooz B Afshar Davoud D

Infection and drug resistance 20200911

<h4>Purpose</h4>Iron is a necessary element for the growth of bacteria; however, there are limited iron sources known for these microorganisms yet. Intracellular iron is stored as ferritin from, which releases iron in a gradual and controlled manner. The present study aimed to characterize ferritin-binding proteins (FBPs) of <i>Streptococcus pneumoniae.</i><h4>Material and methods</h4><i>S. pneumoniae</i> species were cultured in BHI broth containing ferritin (1094 ng/mL) for 4h at 37°C. Ferriti ...[more]

PMID: 32982329

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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

Ferritin Degradation by Pneumococcal HtrA, RadA and ClpP Serine Proteases : A Probable Way For Releasing and Acquisition Of Iron.

Publications

Ferritin Degradation by Pneumococcal HtrA, RadA and ClpP Serine Proteases : A Probable Way For Releasing and Acquisition Of Iron.

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