7 S.D. with 36.3% similarity and 27.1% identity, showing that the two sequences are homologous. Internal five TMS repeats in some 10 TMS transporters In this section, some 10 TMS proteins are shown to have arisen by duplication of a 5 TMS element. A representative putative ten TMS uptake porter, RnsC (TC# 3.A.1.2.12) and its close homologues, IWR-1 molecular weight usually predicted to have a 10 TMS topology using TOPCONS [26], and TMHMM (http://www.cbs.dtu.dk/services/TMHMM/), but predicted to have 8 or 9 TMSs using HMMTOP, takes up ribonucleosides GSK621 nmr and their 2-deoxy derivatives. The topological predictions obtained by the TMHMM program
are shown in Figure 3A. It seemed possible that what appears to be TMSs 1–5 and TMSs 6–10 are repeats. It should be noted, however, that topological predictions by the various programs were not consistent, and that some find more uncertainty exists for this protein and its close homologs. This conclusion did not prevent establishment of the proposed internal repeat.
Figure 3 Internal 5 TMS repeats in some 10 TMS transporters. A (left). Hydropathy plot of RnsC (TC# 3.A.1.2.12). Blue lines denote Hydropathy; Red lines denote Amphipathicity; Orange bars mark transmembrane segments as predicted by HMMTOP. B (right). Putative TMSs 1– 5 of gi222147212 are aligned with putative TMSs 6–10 of gi218884703, yielding a comparison score of 14.9 S.D. with 41.1% similarity and 29.5% identity. The numbers at the beginning of each line refer to the residue numbers in each of the proteins. TMSs are indicated in red lettering. Vertical lines
indicate identities; colons indicate close similarities, and periods indicate more distant similarities. The RnsC protein was NCBI BLASTed to obtain homologues, which were run through CD-Hit to eliminate redundant and strikingly similar sequences (cut off of 80%). The remaining hits were aligned using the ClustalX program. Using SSearch, putative TMSs 1–5 of all homologues were compared with putative TMSs 6–10. The results showed that homologues in GenBank gi222147212 Cytidine deaminase and gi218884703, probably contain internal five TMS duplications (see Additional file 1: Figure S4A and Figure S4B, respectively). When the first half of gi222147212 was aligned with the second half of gi218884703, a comparison score of 14.9 S.D. with 41.1% similarity and 29.5% identity was obtained (Figure 3B). Internal repeats of 5 TMSs in other 10 TMS transporters, and of 10 TMSs in 20 TMS transporters In this section, we examine other putative 10 TMS proteins and compare predictions with 3-dimensional structures. BtuC (TC# 3.A.1.13.1), a vitamin B12 porter constituent, which contains ten TMSs according to the high resolution X-ray crystallographic structure [6], was first examined. However, the WHAT, HMMTOP and TMHMM 2.0 programs all predicted nine TMSs (Figure 4). The topological predictions by WHAT and by X-ray crystallography are shown in Figures 4 and 5, respectively. The missing TMS in Figure 4 is between putative TMSs 7 and 8.