Proteome and Differential Expression Analysis of Membrane and Cytosolic Proteins from Mycobacterium avium subsp. paratuberculosis Strains K-10 and 187.

Authors: Radosevich, Thomas; Reinhardt, Timothy; Lippolis, John; Bannantine, John; Stabel, Judith

Submitted to: Journal of Bacteriology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/15/2006
Publication Date: 2/1/2007
Citation: Radosevich, T.J., Reinhardt, T.A., Lippolis, J.D., Bannantine, J.P., Stabel, J.R. 2007. Proteome and Differential Expression Analysis of Membrane and Cytosolic Proteins from Mycobacterium avium subsp. paratuberculosis Strains K-10 and 187. Journal of Bacteriology. 189(3):1109-1117.

Interpretive Summary: Johne's disease is a chronic, debilitating intestinal disorder in cattle characterized by diarrhea, reduced feed intake, weight loss and death. Cattle usually become infected as young calves by ingesting feces containing the causative bacteria. However, symptoms of disease do not usually present themselves until the animals reach 3 to 5 years of age or even older. During this time the animal is infected and may be shedding the organism in its feces without showing any clinical signs of disease. In addition to reduced milk production by these animals, they also present a potential infective threat to the rest of the herd. Johne’s disease is difficult to diagnose and therefore to control. Development of accurate and sensitive diagnostic tests is dependent upon understanding virulence mechanisms of the bacterium. This paper provides information on a survey of the major classes of proteins that are expressed by the bacterium. This information will be helpful in developing better diagnostic tools for the control and management of paratuberculosis.

Technical Abstract: Little is known of the protein expression in Mycobacterium avium subspecies paratuberculosis (MAP) and how this contributes to pathogenesis. In the present study, proteins from both outer membranes and cytosol were prepared from two strains of MAP; a laboratory-adapted strain K-10 and a recent isolate, strain 187, obtained from a cow exhibiting clinical signs of Johnes disease. SDS-PAGE of cytosol and membrane proteins from K-10 and 187 showed marked differences in protein expression. Relative levels of protein expression from both MAP strains were measured using amine-reactive isobaric tagging reagents (iTRAQ) and tandem mass spectroscopy. Protein identification and relative expression data were obtained for 874 membrane and cytosolic proteins from the MAP proteome. These data showed a number of significant differences in protein expression between K-10 and the clinical isolate 187. Examples of proteins expressed at higher levels in strain 187 as compared to strain K-10 are AtpC, RpoA and several proteins involved in fatty acid biosynthesis. In contrast, proteins such as AhpC and several proteins involved in nitrogen metabolism were expressed at higher levels in strain K-10 as compared to strain 187. These data may provide insights into the proteins whose expression is important in natural infection but are modified once MAP is adapted to laboratory cultivation. Results from these studies will provide tools for developing a better understanding of MAP infection in the host, and offer potential as diagnostic reagents and vaccine candidates.