, 2008; Collin et al., 2011). Accessory proteins can stabilize the secretin itself, the secretin subunits prior to membrane insertion or are co-dependent with the secretin for mutual stability. Accessory Forskolin proteins are membrane-associated and contain periplasmic regions that are thought to interact directly with the secretin. Systems may contain either a pilotin, an accessory protein(s), or both. Conservation of particular genes across a system does not necessary correlate with similar function, as significant differences

have been documented between bacterial species. Pilotins that have been identified and characterized to date are listed in Table 1. Although many systems have identifiable pilotin orthologues, they are either absent or have yet to be identified in others. Competence systems, filamentous phage, and T4bP each lacks pilotins. Most T2S systems characterized to date have pilotins, except for Pseudomonas aeruginosa Hxc and Xcp, Escherichia coli Gsp, Aeromonas hydrophila Exe, and Vibrio cholerae Eps. Immediate PD-0332991 cell line differences can be found between the remaining pilotin-containing T2S, T3S, and T4aP systems by comparing the genomic organization of the genes encoding the secretin subunit and the pilotin. The gene encoding the secretin subunit is typically clustered with other genes that encode a variable number of proteins involved in

system assembly. The T2S pilotins Erwinia chrysanthemi outS and Klebsiella oxytoca pulS as well as the T3S pilotins

Salmonella typhimurium invH, Shigella flexneri Ergoloid mxiM and Yersinia enterocolitica yscW are each encoded with other components of their respective assembly systems. In contrast, the T4aP pilotins Pseudomonas pilF (pilW), Neisseria pilW, and Myxococcus xanthus tgl are located elsewhere in the genome and surrounded by non-T4aP genes. While pilotins fulfill similar roles in localizing and/or assembling the secretin, the structure of specific pilotins can vary significantly. Based on the available structural data or on sequence-based predictions, we divided pilotins into one of three different classes: Class 1 pilotins are composed entirely of α-helical tetratricopeptide repeats (TPRs) and are roughly double the size of other pilotins. Class 2 pilotins are comprised predominantly of β-strands, while Class 3 pilotins are predominantly α-helical non-TPR proteins. The structure of pilotins clearly divides the secretion and pilus systems. Sequence identity among T4aP pilotins PilF, PilW, and Tgl is poor, ranging from 13% to 25%. However, the structures of PilF and PilW that have been determined by X-ray crystallography (Koo et al., 2008; Trindade et al., 2008) show that they have a common protein fold. PilF and PilW are each composed of six TPRs with a nearly identical tertiary fold (Fig. 1a).