Isolation and 2-D-DIGE proteomic analysis of intracellular and extracellular forms of Listeria monocytogenes

The pathogenicity of Listeria monocytogenes is related to its ability of invading and multiplying in eukaryotic cells. Its main virulence factors are now well characterized, but limited proteomic data is available concerning its adaptation to the intracellular environment. In this study, L. monocyto...

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Bibliographic Details
Published in:Proteomics (Weinheim) 2009-12, Vol.9 (24), p.5484-5496
Main Authors: Van de Velde, Sébastien, Delaive, Edouard, Dieu, Marc, Carryn, Stéphane, Van Bambeke, Françoise, Devreese, Bart, Raes, Martine, Tulkens, Paul M
Format: Article
Language:English
Subjects:
2-D-DIGE
Amino Acids - metabolism
Analytical, structural and metabolic biochemistry
Bacterial Proteins - analysis
Bacterial Proteins - metabolism
Biological and medical sciences
Carbohydrate Metabolism
Cell Line
Electrophoresis, Gel, Two-Dimensional
Fundamental and applied biological sciences. Psychology
Humans
Intracellular
Listeria monocytogenes
Listeria monocytogenes - isolation & purification
Listeria monocytogenes - metabolism
Listeriosis - microbiology
Microbiology
Miscellaneous
Monocytes - microbiology
Nucleotides - metabolism
Peptide Synthases - analysis
Peptide Synthases - metabolism
Proteins
Proteome - analysis
Proteome - metabolism
Proteomics
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Thiamine - metabolism
THP-1 monocytes
Viral Envelope Proteins - analysis
Viral Envelope Proteins - metabolism
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Summary:The pathogenicity of Listeria monocytogenes is related to its ability of invading and multiplying in eukaryotic cells. Its main virulence factors are now well characterized, but limited proteomic data is available concerning its adaptation to the intracellular environment. In this study, L. monocytogenes EGD (serotype 1/2a) grown in human THP-1 monocytes (24 h) were successfully separated from host organelles and cytosolic proteins by differential and isopycnic centrifugation. For control, we used cell homogenates spiked with bacteria grown in broth. Proteomes from both forms of bacteria were compared using a 2-D-DIGE approach followed by MALDI-TOF analysis to identify proteins. From 1684 distinct spots, 448 were identified corresponding to 245 distinct proteins with no apparent contamination of host proteins. Amongst them, 61 show underexpression (stress defense; transport systems, carbon metabolism, pyrimidines synthesis, D-Ala-D-Ala ligase) and 22 an overexpression (enzymes involved in the synthesis of cell envelope lipids, glyceraldehyde-3-phosphate, pyruvate and fatty acids). Our proteomic analysis of intracellular L. monocytogenes (i) suggests that bacteria thrive in a more favorable environment than extracellularly, (ii) supports the concept of metabolic adaptation of bacteria to intracellular environment and (iii) may be at the basis of improved anti-Listeria therapy.
ISSN:1615-9853
1615-9861
1615-9861
DOI:10.1002/pmic.200900503