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Published on: March 17, 2019
Proteome analysis of Madrid E strain of Rickettsia prowazekii
Chien-Chung Chao1, Dirk Chelius, Terry Zhang
1Rickettsial Diseases Department, Infectious Diseases Directorate, Naval Medical Research Center, Silver Spring, MD 20910-7500, USA. chaoc@nmrc.navy.mil
Abstract:
Rickettsia prowazekii, an obligate intracellular Gram-negative bacterium, is the etiologic agent of epidemic typhus. The threat of typhus as a biological weapon lies in its stability in the dried louse feces and in its infection by inhalation of an aerosol. Consequently, it is listed as a select agent and warrants more research to understand its pathogenesis. Although the genomic DNA sequence of strain Madrid E has been completed, the actual expression of the individual protein has not been investigated. In order to provide a global view of the expressed protein profile, the whole cell lysate of purified rickettsia (Madrid E strain) was reduced, alkylated, and digested with trypsin. The total digest was characterized by a two-dimensional liquid chromatography mass spectrometry system and analyzed with a modified version of the ProteomeX workstation. A total of 252 proteins out of 834 predicted protein-coding genes were identified, 238 proteins were identified by the detection of at least two unique peptides. Only 14 proteins were identified by the detection of one unique peptide in all three separate analyses. Among the 238 proteins identified by multiple unique peptides, 230 proteins were found in at least two of three separate analyses. The reproducible and convenient methodology and the information described here have provided a foundation for future proteome study of various R. prowazekii strains with different virulence.
Insights
This study identifies 252 proteins expressed by Rickettsia prowazekii (Madrid E strain), providing a foundational proteome map. This research enhances understanding of epidemic typhus pathogenesis and aids future virulence studies.
Area of Science:
- Microbiology
- Proteomics
- Bacteriology
Background:
- Rickettsia prowazekii causes epidemic typhus and is a potential biological weapon due to its stability and aerosol transmission.
- Understanding R. prowazekii pathogenesis requires knowledge of its expressed proteins, beyond its sequenced genome.
Purpose of the Study:
- To globally characterize the proteome of Rickettsia prowazekii (Madrid E strain).
- To establish a reproducible methodology for future R. prowazekii proteomic studies.
Main Methods:
- Purified R. prowazekii (Madrid E strain) whole cell lysate was processed (reduced, alkylated, trypsin digested).
- Samples were analyzed using two-dimensional liquid chromatography coupled with mass spectrometry (2D-LC-MS/MS).
- Data was analyzed using a modified ProteomeX workstation.
Main Results:
- A total of 252 proteins were identified from 834 predicted protein-coding genes.
- 238 proteins were identified by at least two unique peptides, with 230 found in at least two of three analyses.
- 14 proteins were identified by a single unique peptide across all analyses.
Conclusions:
- The study provides a comprehensive proteomic profile for R. prowazekii (Madrid E strain).
- The developed methodology is reproducible and suitable for comparative proteomic studies of different R. prowazekii strains.
- This proteomic data serves as a basis for further research into R. prowazekii virulence mechanisms.
