J Vet Sci.  2018 Mar;19(2):216-231. 10.4142/jvs.2018.19.2.216.

Analysis of protein expression in Brucella abortus mutants with different growth rates by two-dimensional gel electrophoresis and LC-MS/MS peptide analysis

• Park WB
• Im YB
• Shim S
• Yoo HS

• KMID: 2407621
• DOI: http://doi.org/10.4142/jvs.2018.19.2.216

Abstract

Brucella abortus is a bacterium that causes brucellosis and is the causative agent of worldwide zoonoses. Pathogenesis of the B. abortus infection is complicated, and several researchers have attempted to elucidate the infection mechanism of B. abortus. While several proteins have been revealed as pathogenic factors by previous researchers, the underlying mechanism of B. abortus infection is unresolved. In this study, we identified proteins showing different expression levels in B. abortus mutants with different biological characteristics that were generated by random insertion of a transposon. Five mutants were selected based on biological characteristics, in particular, their growth features. Total proteins of mutant and wild-type B. abortus were purified and subjected to two-dimensional gel electrophoresis. Thirty protein spots of each mutant with expression increases or decreases were selected; those with a change of more than 2-fold were compared with the wild-type. Selected spots underwent liquid chromatography tandem mass spectrometry for peptide analysis. DnaK and ClpB, involved in protein aggregation, increased. SecA and GAPDH, associated with energy metabolism, decreased in some mutants with a growth rate slower than that of the wild-type. Mutants with slower growth showed a decrease in energy metabolism-related proteins, while mutants with faster growth showed an increase in pathogenicity-related proteins.

Keyword

Brucella abortus; growth rates; protein sequence analysis; two-dimensional gel electrophoresis

MeSH Terms

Brucella abortus*
Brucella*
Brucellosis
Chromatography, Liquid
Electrophoresis, Gel, Two-Dimensional*
Energy Metabolism
Population Characteristics
Sequence Analysis, Protein
Tandem Mass Spectrometry
Zoonoses

Figure

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  Fig. 1 Two-dimensional gel electrophoresis (2-DE) image analysis of Brucella abortus wild-type and B. abortus mutant C3. (A) 2-DE image of B. abortus wild-type. (B) 2-DE image of B. abortus mutant C3. (C) Spots of B. abortus mutant C3 that were relatively increased compared to B. abortus wild-type. (D) Spots of B. abortus mutant C3 that were relatively decreased compared to B. abortus wild-type.

• 

  Fig. 2 Two-dimensional gel electrophoresis (2-DE) image analysis of Brucella abortus wild-type and mutant C8. (A) 2-DE image of B. abortus wild-type. (B) 2-DE image of B. abortus mutant C8. (C) Spots of B. abortus mutant C8 that were relatively increased compared to B. abortus wild-type. (D) Spots of B. abortus mutant C8 that were relatively decreased compared to B. abortus wild-type.

• 

  Fig. 3 Two-dimensional gel electrophoresis (2-DE) image analysis of Brucella abortus wild-type and mutant C13. (A) 2-DE image of B. abortus wild-type. (B) 2-DE image of B. abortus mutant C13. (C) Spots of B. abortus mutant C13 that were relatively increased compared to B. abortus wild-type. (D) Spots of B. abortus mutant C13 that were relatively decreased compared to B. abortus wild-type.

• 

  Fig. 4 Two-dimensional gel electrophoresis (2-DE) image analysis of Brucella abortus wild-type and mutant C24. (A) 2-DE image of B. abortus wild-type. (B) 2-DE image of B. abortus mutant C24. (C) Spots of B. abortus mutant C24 that were relatively increased compared to B. abortus wild-type. (D) Spots of B. abortus mutant C24 that were relatively decreased compared to B. abortus wild-type.

• 

  Fig. 5 Two-dimensional gel electrophoresis (2-DE) image analysis of Brucella abortus wild-type and mutant C30. (A) 2-DE image of B. abortus wild-type. (B) 2-DE image of B. abortus mutant C30. (C) Spots of B. abortus mutant C30 that were relatively increased compared to B. abortus wild-type. (D) Spots of B. abortus mutant C30 that were relatively decreased compared to B. abortus wild-type.

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