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Periplasmic Proteome of Yersinia pestis KIM 6+

Profiling the Periplasmic Proteome of Yersinia pestis, strain KIM6+

A comprehensive analysis of the periplasmic proteome of Yersinia pestis strain KIM6+ was performed. The KIM6+ strain of the plague bacterium Y. pestis is an avirulent derivative of the fully virulent KIM strain, which was cured of the pCD1 plasmid. The chromosomal pgm locus and the plasmids pMT1 and pPCP1 are present in KIM6+ [1]. Periplasmic fractions were isolated using the spheroplasting method by treating cells with lysozyme and EDTA that results in cell wall degradation and release of macromolecules present in the periplasmic space [2]. Low-level lysis of spheroplasts caused experimental contamination with cytoplasmic proteins. To discern true periplasmic from cytoplasmic contaminant proteins, differential 2-DE display comparing protein profiles of the periplasmic with a cytoplasmic fraction was applied. Proteins enriched in the periplasmic fraction were assigned to the periplasm (for further information, see Table). In addition, Mr and pI values of protein spots in 2-DE gels and sequence coverage at the N-termini of proteins were assessed to evaluate the absence of signal peptides in mature periplasmic proteins. This data was also compared with bioinformatically predicted signal peptide cleavage sites (for further information, see Table). In addition to all proteins experimentally assigned to the periplasm of Y. pestis (161 entries), ca. 50 cytoplasmic proteins are also displayed in the 2-DE gel profiles in Figures 1-3 and described in the Table. Spot numbers denoted in Figures 1-3 are equivalent to spot numbers listed in column 1 of the Table. In the Table, annotation data of proteins (KIM strain, protein database in NCBI [3]) and additional data describing protein traits (functions, conserved domains, subcellular assignments) are provided.

1. Fetherston, J.D. and R.D. Perry, The pigmentation locus of Yersinia pestis KIM6+ is flanked by an insertion sequence and includes the structural genes for pesticin sensitivity and HMWP2. Mol Microbiol, 1994. 13(4): p. 697-708.
2. Lucier, T.S., et al., Iron uptake and iron-repressible polypeptides in Yersinia pestis. Infect Immun, 1996. 64(8): p. 3023-31.
3. Deng, W., et al., Genome sequence of Yersinia pestis KIM. J Bacteriol, 2002. 184(16): p. 4601-11.

Figures

Figure 1A

Figure 1B

Figure 2

Figure 3A

Figure 3B

Figure 3C

Tables