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Figure 1
Design strategy for the Tako and Ika proteins. PDB entry 2ovp was identified as a pseudo-symmetrical protein consisting of eight repeats arranged around a central axis (a). The repeats were superposed (b). The multiple sequence alignment (c) and corresponding guide tree (d) indicate that the eighth repeat differs from the others. Each repeat from repeat 2 to repeat 6 was used individually to create perfectly symmetrical propeller backbones. The second repeat yielded the backbone with the lowest energy (e). The ancestral sequences, created from the sequence alignment and corresponding guide tree, were then mapped onto this backbone and the structure was optimized using PyRosetta. The sequence with the lowest r.m.s.d. (blue dot) from the starting backbone was chosen as the Tako8 protein (f). The crystallographically determined Tako8 structure (g) proved to be essentially identical to the design model. Overlays of the experimental models (rainbow-coloured) and designs (white) are shown for Tako8 (h) and Ika8 (i). An alignment between the repeats of Tako8, the alternating repeats of Ika and the two previous attempts at designing a perfectly symmetric WD40 protein shows the differences that may contribute to the stability of the protein (j). The highly conserved Trp–Ser–His–Asp motif commonly observed in WD40 proteins is shown by red arrows above the sequence alignment.

ISSN: 2052-2525