trachomatis Inc proteins. To know whether this result also applied to C. pneumo niae, selleck chem inhibitor we raised antibodies against 7 putative Inc proteins from C. pneumoniae as GST tagged fusion proteins. As a control we used antibodies against the C. pneumoniae Inc protein CPn0186. The anti fusion protein antibodies were used to localize the endogenous proteins in cells infected by C. pneumoniae for 96 hours. In contrast to the inclusion labeling observed with anti CPn0186 antibodies, none of the 7 sera stained the inclusion membrane. The detection of endogenous antigens was removed by pre absorption with corresponding GST fusion proteins but not heterologous Inhibitors,Modulators,Libraries GST fusion proteins, demonstrating the specificity of the antibodies.
While they did not stain the inclusion membrane, the 7 sera labeled the bacteria, demonstrating that the corresponding proteins Inhibitors,Modulators,Libraries are expressed at this stage of infection, and remain bac teria associated. We cannot exclude the possibility that some or all of these proteins Inhibitors,Modulators,Libraries are partially exposed on the membrane and not detected by this approach. How ever, we can conclude that these 7 putative Inc proteins are not constitutively secreted. Table 4 recapitulates the list of putative Inc proteins for these two species with the TTS and localization data. Discussion and Conclusions Initially, to identify all putative Inc proteins, we started from the IncA domain from Pfam database, which is derived from the multiple alignment of IncA like sequences. This domain includes the hydrophobic domain and an adjacent coiled coil region, which are characteristics of IncA.
When used to detect Inc pro teins, this model misclassified Inc proteins sequences which are devoid of coiled coil regions and appeared far down in noise rank, with a Inhibitors,Modulators,Libraries non significant Score Evalue. This indicates that the Pfam IncA domain is too specific for a large scale genomic analysis. Known Inc proteins contain two transmem brane alpha helical segments separated by a loop of less than 30 amino acids. Using this criteria and bioinfor matics tools, we have searched for all putative Inc pro teins in seven chlamydial proteomes and obtained 537 candidates. These results were validated experimentally for C. trachomatis and C. pneumoniae, as we found that Inhibitors,Modulators,Libraries 90% of the putative Inc proteins of these species had a TTS signal, which is a property of Inc proteins indepen dent of their hydrophobicity profile. Secondary structure analysis revealed that Inc proteins are enriched in coiled coil domains. In bacteria, coiled coil www.selleckchem.com/products/Lenalidomide.html containing proteins represent 5% of proteins, and the majority contain only one helix of around 28 residues. Extended coiled coil domains are rare and are enriched in type III secretion proteins.