Average hydropathy plot (dark red line) showing the basis for the topological predictions made for the Nectria haematococca (Fusarium solani) nhTMEM16 (anoctamin) protein (TC: 1.A.17.1.18), for which x-ray structures are available (PDB IDs 4WIS and 4WIT).

Vertical tan bars show the positions of the predicted TMSs using the Loop Finder program (V. S. Reddy and M. H. Saier, unpublished). The green bar shows the position of the α-helix corresponding to TMS 6. This helix was not predicted to be a TMS by this program, HMMTOP [96] or CCTOP [108], although the x-ray structure confirmed that it is one. HMMTOP predicted TMSs 1 and 2 as a single TMS, although the structure confirms that the corresponding hydrophobicity peak is composed of two TMSs. The two purple bars, representing the position of transmembrane helices 7 and 8 in the x-ray structure, were predicted by these programs and AveHAS [106] to be a single TMS (also note the 7^th hydrophobicity peak in Fig 4A). This explains the discrepancy in the predictions for different members of the Anoctamin Superfamily (between 8 and 10 TMSs). The locations, in the hydropathy curve, of the three pairs of functional residues that bind Ca²⁺ in TMSs 6, 7 and 8 are depicted with blue, black and green circles, respectively.