Mapping phosphoproteins in Mycoplasma genitalium and Mycoplasma pneumoniae

Su, Hsun-Cheng; Hutchison, Clyde A.; Giddings, Morgan C.

doi:10.1186/1471-2180-7-63

Cited by 42 publications

(35 citation statements)

References 58 publications

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“…Several of the proteins that are involved in cytadherence in M. pneumoniae are very large proteins, such as HMW1, HMW2, and HMW3. It has been shown that these proteins are phosphorylated in M. pneumoniae (24,33). Therefore, we decided to analyze specifically the phosphorylation pattern of large proteins by using 6% SDS-polyacrylamide gels.…”

Section: Resultsmentioning

confidence: 99%

“…Among these proteins are HMW1 and HMW2, large cytadherence proteins that were shown to be phosphorylated on serine and threonine residues (9,24). Recently, a proteomic approach was used to identify phosphorylated proteins in M. genitalium and M. pneumoniae (33). This study identified 18 phosphoproteins in M. pneumoniae, among them the surface protein MPN474, the cytadherence protein HMW3, and several metabolic enzymes.…”

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confidence: 99%

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The Stability of Cytadherence Proteins in Mycoplasma pneumoniae Requires Activity of the Protein Kinase PrkC

et al. 2010

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Mycoplasma pneumoniae belongs to the mollicutes, a group of bacteria that have strongly reduced genomes but that are nevertheless capable of independent life. With only three transcription factors, the regulatory features of these bacteria are very limited. Thus, posttranslational regulation might be important for M. pneumoniae. In addition to the highly specific HPr kinase, the M. pneumoniae prkC gene encodes the serine/ threonine protein kinase C. In order to study the function(s) of this kinase, we isolated an M. pneumoniae mutant affected in PrkC. This mutation resulted in nonadherent growth and loss of cytotoxicity. Examination of the phosphorylation profile of the prkC mutant suggested that phosphorylation of cytadherence proteins was affected by the loss of this kinase. In contrast, inactivation of the prpC gene affecting the protein phosphatase that antagonizes PrkC-dependent phosphorylation resulted in more intensive phosphorylation of the cytadherence proteins HMW1 and HMW3 of the major adhesin P1 and of the surface protein MPN474. Moreover, loss of PrkC affects not only the phosphorylation state of the cytadherence proteins but also their intracellular accumulation. However, the expression of the corresponding genes was not affected by PrkC, suggesting that PrkC-dependent phosphorylation results in stabilization of the cytadherence proteins. The HMW proteins and P1 are part of the so-called terminal organelle of M. pneumoniae that is involved in gliding motility, cell division, and adhesion to host epithelial tissues. Our observations suggest that the posttranslational modification of cytadherence proteins by PrkC is essential for the development and function of the M. pneumoniae terminal organelle.Mycoplasma pneumoniae belongs to the mollicutes, i.e., cell wall-less bacteria. These organisms are among the smallest selfreplicating living beings capable of a host-independent existence. M. pneumoniae is a pathogenic bacterium that causes atypical pneumonia and extrapulmonary infections, such as autoimmune disorders, asthma, and arthritis (4,32,34).M. pneumoniae and its close relative Mycoplasma genitalium have recently attracted much attention, not only with respect to the elucidation of pathogenicity mechanisms but also because the small genomes of these bacteria define the lower limit of naturally existing independent life. The analysis of the minimal gene complement of M. pneumoniae and M. genitalium is one of the sources of synthetic biology, a new discipline of biology (12, 13).However, life is not static and not determined only by a defined set of genes. A very important feature is the control of biological activities in response to changing environmental conditions. Only this control enables organisms to adapt to different habitats and to survive suboptimal conditions. In M. pneumoniae, the regulatory potential seems to be rather limited. In bacteria, regulation of gene expression is achieved mainly at the level of transcription, by alternative sigma factors of the RNA polymerase, by transcriptio...

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Section: Resultsmentioning

confidence: 99%

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confidence: 99%

The Stability of Cytadherence Proteins in Mycoplasma pneumoniae Requires Activity of the Protein Kinase PrkC

et al. 2010

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“…Phosphoproteins separated by gel electrophoresis can also be visualized by the use of a phosphospecific stain such as Pro-Q Diamond [16][17][18][19]. Alternatively, phosphoproteins can be radioactively labeled using 32 P or 33 P [16,17] and detected by autoradiography, which is the most sensitive method available, but may not be compatible with downstream analytical procedures such as MS. Radioactive labeling is possible both in vivo as well as in vitro. In in vivo studies, cells are incubated with 32 P, however, the presence of endogenous ATP pools within cells can interfere with the incorporation of the label and result in inefficient radioactive labeling [20].…”

Section: Detection Of Phosphoproteinsmentioning

confidence: 99%

Analytical strategies for phosphoproteomics

2009

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Protein phosphorylation is a key regulator of cellular signaling pathways. It is involved in most cellular events in which the complex interplay between protein kinases and protein phosphatases strictly controls biological processes such as proliferation, differentiation, and apoptosis. Defective or altered signaling pathways often result in abnormalities leading to various diseases, emphasizing the importance of understanding protein phosphorylation. Phosphorylation is a transient modification, and phosphoproteins are often very low abundant. Consequently, phosphoproteome analysis requires highly sensitive and specific strategies. Today, most phosphoproteomic studies are conducted by mass spectrometric strategies in combination with phosphospecific enrichment methods. This review presents an overview of different analytical strategies for the characterization of phosphoproteins. Emphasis will be on the affinity methods utilized specifically for phosphoprotein and phosphopeptide enrichment prior to MS analysis, and on recent applications of these methods in cell biological applications.

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“…The minimum set of genes, also called essential genes, in both prokaryotes and eukaryotes, are those described as indispensable for the survival of an organism and are therefore the basis of life for a particular organism. 25 identified only 480 protein-coding regions, while a later study by Ueno et al (2008) 3 found 484 coding regions. These identified coding regions include genes for DNA replication, transcription, translation, DNA repair, cellular transport and energy metabolism.…”

Section: 21mentioning

confidence: 92%