fPOP: footprinting functional pockets of proteins by comparative spatial patterns

Abstract

fPOP (footprinting Pockets Of Proteins, http://pocket.uchicago.edu/fpop/) is a relational database of the protein functional surfaces identified by analyzing the shapes of binding sites in approximately 42,700 structures, including both holo and apo forms. We previously used a purely geometric method to extract the spatial patterns of functional surfaces (split pockets) in approximately 19,000 bound structures and constructed a database, SplitPocket (http://pocket.uchicago.edu/). These functional surfaces are now used as spatial templates to predict the binding surfaces of unbound structures. To conduct a shape comparison, we use the Smith-Waterman algorithm to footprint an unbound pocket fragment with those of the functional surfaces in SplitPocket. The pairwise alignment of the unbound and bound pocket fragments is used to evaluate the local structural similarity via geometric matching. The final results of our large-scale computation, including approximately 90,000 identified or predicted functional surfaces, are stored in fPOP. This database provides an easily accessible resource for studying functional surfaces, assessing conformational changes between bound and unbound forms and analyzing functional divergence. Moreover, it may facilitate the exploration of the physicochemical textures of molecules and the inference of protein function. Finally, our approach provides a framework for classification of proteins into families on the basis of their functional surfaces.

Figure 1.

Illustration of the fPOP shape analysis. (a) Identification of a split pocket in a bound structure as a spatial template (a collection of 38 900 spatial templates). (b) Surface segmentation of an unbound form. (c) Geometrically matching the spatial pattern of the template with those of putative pockets in the unbound form. (d) Measuring features and footprinting the binding surface of an unbound form.

Figure 2.

Predicting the binding surfaces of unbound forms. (a) The binding surface (the 13th pocket colored green with a mouth colored blue) of the galactose-binding protein (pdb1gcg) has a spatial pattern footprinted by the 16 functional surfaces of the 14 similarity hits in SplitPocket. (b) The functional surface (pdb3b6u.B) of a human motor protein is distantly related to that of the galactose-binding protein. A binding-ligand ADP (red) interacts with the split pocket (green). (c) The binding surface (the 11th pocket) of the triose phosphate isomerase (pdb1ypi.A) is correctly predicted. The fPOP shape analysis indicates that significant local conformational changes (4.1 Å RMSD) occur between the apo-form (pdb 1ypi.A) and the holo-form (pdb2ypi) in (d).

Figure 3.

Footprinting the binding surface of a tyrosine kinase by a remote homologous protein. (a) At a significant RMSD P-value of 4 × 10⁻⁷, the binding surface (green) of pdb1yoj.A is matched with the binding pocket of pdb3c4w.A split by an ATP (red). (b) The optimal alignment of the binding surfaces between the query (pdb1yoj.A, red) and a spatial template (pdb3c4w.A, black) is used to compute their shape similarity at a RMSD of 2.3 Å. The similarity of pocket-fragments (43%) is considerably higher than that of the full-length primary sequences (22.3%). The catalytic residues (R³⁹⁰, A³⁹² and N³⁹³) of pdb1yoj.A are also aligned with those (K³¹⁶, E³¹⁸ and N³¹⁹) of pdb3c4w.A.

Figure 4.

A structural phylogeny of binding surfaces for a subset of ATP-binding kinases.

Figure 5.

Characterization of the functional surface of an alpha-amylase (pdb1bag). (a) The 19th pocket (green) is split by glucose (red). (b) The mouth of the split pocket has a hydrophobic accessible area (blue, 165.4 Ų). (c) The highest SCI (0.898) occurs in the split pocket. The spatial pattern of this functional surface consists of 19 residues with conservation weights for assessing the evolutionary characteristics. Four catalytic residues D¹⁷⁶, H¹⁸⁰, Q²⁰⁸ and D²⁶⁹ are highly conserved. In addition, there are 10 important residues sitting on the mouth. Among them, seven are hydrophobic residues indicated by asterisk.