Exocyst Helical Bundles Figure 1. The exocyst subunits have similar helical bundle structures. (A) The known structures of the exocyst subunits are shown: Exo70p (PDB ID 2B1E), Exo84CT (PDB ID 2D2S), Sec15CT (PDB ID 2A2F), Sec6CT (PDB ID 2FJI). Molecular graphics were generated with PyMOL (http://pymol. sourceforge.net/). Exo84CT is aligned with the N-terminal helical bundles of Exo70p, while Sec15CT and Sec6CT are aligned with the C-terminal bundles of Exo70p. (B) Secondary structure predictions for all of the exocyst subunits. The black horizontal lines represent the sequence of each yeast exocyst subunit. The predicted a-helices (magenta) and b-strands (cyan) are indicated by vertical bars above each line. The height of the bars is proportional to the confidence of the secondary structure prediction [41]. Red blocks underline regions of the known structures. Green blocks underline the best hits to exocyst structures (see Table 1). doi:10.1371/journal.pone.0004443.g001 slightly soluble protein. An alternative approach is to computa- tionally predict domain structures based on the similarity to proteins with known structural domains [20,22–24]. However, this approach is challenging if the protein has little or no similarity with proteins of known structures. In that case, more sensitive computational methods, such as hidden Markov model (HMM) predictions [25], may be successful. Based on the structural conservation observed in the exocyst subunits, we hypothesized that the other subunits would have similar helical bundle structures [3]. Therefore, we examined the conservation of structural similarity between the subunits by profile HMM analyses using the HHSearch program [26]. Profile HMMs are similar to simpler sequence profiles, but in addition to the amino acid frequencies in the columns of a multiple sequence alignment, they contain information about the frequency of inserts and deletions at each column, plus transition probabilities. In addition, secondary structure can be included in the HMM-HMM comparison, leading to another increase in sensitivity. We applied state-of-the-art HMM-HMM comparisons to the exocyst complex and detected structural similarity between all of the exocyst subunits. We verified these structure predictions by identifying a structural domain in one of the exocyst subunits, Sec10p (YLR166C), from Saccharomyces cerevisiae. Sec10p is one of the core subunits in the complex and has previously been shown to interact in vitro with its partner exocyst subunits Sec6p, Exo70p and Sec15p PLoS ONE | www.plosone.org 2 February 2009 | Volume 4 | Issue 2 | e4443