Article Title: The Integrative Conjugative Element (ICE) of Mycoplasma agalactiae: Key Elements Involved in Horizontal Dissemination and Influence of Coresident ICEs
Figure Lengend Snippet: Overview of conjugative ICE transfer in M. agalactiae ). Under normal conditions, ICEA copies are found integrated into the host chromosome and most ICEA genes are not expressed. Among the few proteins expressed by chromosomal ICEAs is the CDS14 lipoprotein, which is surface exposed and plays a critical role in initiating the conjugative process (step 1). When ICEA gene expression is induced, under specific cellular conditions or stochastically, the cis -acting DDE transposase is produced and one of the three ICEA copies excises from the chromosome and forms a circular double-stranded DNA (dsDNA) molecule (step 2). ICEA circularization induces the expression of the conjugative module, whose products assemble into the mating pore, a simplified form of type IV secretion system (T4SS) found in more-complex bacteria (step 3). A protein complex known as a relaxosome recognizes the origin of transfer ( oriT ) on the circular ICEA, and a relaxase generates a linear single-stranded DNA (ssDNA) by nicking the ICEA DNA (step 4). Finally, the relaxosome complex interacts with the TraG-like (VirD4 homologue) energetic component found at the inner side of the membrane that facilitates the transfer of the ssDNA bound to the relaxase through the mating channel (step 5). Once in the recipient strain, the ICEA recircularizes, becomes doubly stranded, and integrates randomly into the host chromosome. The minimal functional ICEA encompasses 80% of the coding sequence and includes a gene cluster ( cds5 to cds19 , encoding proteins with transmembrane domains) that most likely represents a module associated with the conjugative channel. Additional essential ICEA determinants included the CDS14 surface lipoprotein, the CDSG putative partitioning protein, and the DDE transposase (CDS22), together with several proteins of unknown function (CDS1, CDSA, CDSC, and CDS30).
Article Snippet: The anti-CDS14 lipoprotein rabbit serum was produced by animal immunization with a recombinant CDS14 protein (pMAL protein fusion and purification system; New England Biolabs).
Techniques: Expressing, Produced, Functional Assay, Sequencing