Abstract
Mycoplasmas have proven to be successful pathogens despite their minimal genomes in some species. Their simplistic nature makes them an important tool for understanding the genes involved in basic metabolic processes and pathogenesis. Of particular interest are the genes involved in mycoplasma evasion of host immune responses and survival within phagocytic cells. The proteins secreted by mycoplasmas in the acidic and hydrolytic environment of a macrophage may help to uncover important virulence factors of this group of prokaryotes. This study examined the effect of pH on the secretome of Mycoplasma capricolum. The bacterium was cultured in protein-free media and the culture supernatant was investigated for proteins produced during growth. Twodimensional gel electrophoresis followed by MALDI-TOF mass spectrometry of tryptic digests was used to identify the proteins in this fraction. Under acidic conditions, a total of a 111 protein spots from the secretome of M. capricolum were visualized by silver staining. A lower number of spots were detected in the neutral culture supernatant (82 spots). Superimposition of the two 2D maps revealed 26 proteins unique to the secretome at acidic pH but identification of all spots was not possible. The identification of neutral proteins revealed the presence of 7 putative lipoproteins, zinc metalloprotease, three peptidases, inorganic pyrophosphatase, nicotinamidase/pyrazinamidase, and ribosomal protein S4, and a number of hypothetical proteins. Proteins identified in the acidic culture supernatant included hemolysin A, peptide methionine sulfoxide reductases, two peptidases, nucleoside kinase, one lipoprotein, and others. An API ZYM enzyme assay demonstrated that the spectrum of enzymes secreted at an acidic pH was similar to that at neutral pH, the activities of the acid phosphatase, the phosphohydrolase, the C8 esterase lipase and the alkaline phosphatase were enhanced. The three key secreted proteins prominent at acidic pH were the acid phosphatase, the hemolysin, and the gelatinase.
Highlights
Members of the mycoplasmas are known to have some the smallest free-living prokaryotes
This study examined the effect of pH on the secretome of Mycoplasma capricolum
Considerable advances have been made towards understanding the pathogenesis of mycoplasmas; of particular interest is the interaction of these pathogens with the immune system, macrophage activation, cytokine induction, superantigens, autoimmune manifestations, and the evasion of host immune responses
Summary
Members of the mycoplasmas are known to have some the smallest free-living prokaryotes. It is suggested that these mollicutes are phylogenetically related to Gram-positive bacteria whereby their genomes were reduced through multiple events of mutations This implies that these simple bacteria are not found at the root of the phylogenetic tree but are more likely late evolutionary products [4]. Mycoplasmas enter their host and multiply over long periods of time, slowly wearing down the host [1] This is one reason perhaps why mycoplasmas have been linked to AIDS pathogenesis and to malignant transformation, chromosomal aberrations, the Gulf War Syndrome, and other unexplained and complex illnesses, including chronic fatigue syndrome, Crohn’s disease, and various arthritis’ [5]. One of the most important questions about the pathogenesis of mycoplasmas is how they cause damage to the host cells and to what extent is the damage clinically apparent in the host
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