Anchor residues in protein-protein interactions

Abstract

We show that the mechanism for molecular recognition requires one of the interacting proteins, usually the smaller of the two, to anchor a specific side chain in a structurally constrained binding groove of the other protein, providing a steric constraint that helps to stabilize a native-like bound intermediate. We identify the anchor residues in 39 protein-protein complexes and verify that, even in the absence of their interacting partners, the anchor side chains are found in conformations similar to those observed in the bound complex. These ready-made recognition motifs correspond to surface side chains that bury the largest solvent-accessible surface area after forming the complex (> or =100 A2). The existence of such anchors implies that binding pathways can avoid kinetically costly structural rearrangements at the core of the binding interface, allowing for a relatively smooth recognition process. Once anchors are docked, an induced fit process further contributes to forming the final high-affinity complex. This later stage involves flexible (solvent-exposed) side chains that latch to the encounter complex in the periphery of the binding pocket. Our results suggest that the evolutionary conservation of anchor side chains applies to the actual structure that these residues assume before the encounter complex and not just to their loci. Implications for protein docking are also discussed.

Fig. 1.

Side-chain dynamics. (A) The rmsd of Arg-39 of ribonuclease A with respect to the structure found in the complex (1DFJ) and the unbound ligand (7RSA). The rmsd was computed for 2,000 snapshots of a 4-ns MD simulation of 7RSA. (B) Clustering distribution of the conformations of Arg-39 (solid line). The top 10 clusters were derived from a pairwise rmsd analysis of the MD snapshots, by using a clustering radius of 2 Å. Bars indicate the rmsd (left vertical axis) of the side chain in the cluster center with respect to the bound (dark-blue bar) and unbound (pink bar) conformations. (Inset) Cluster centers for the largest clusters as well as the bound (blue), unbound (red), and dominant MD (green) conformations. Note that there is no significant sampling of the unbound rotamer.

Fig. 2.

Anchor residues in six complexes. Simulated proteins are shown in cartoon form, and the receptor is shown as surface except for the 1DFJ complex in E. Each anchor side chain is shown in stick conformations that represent the crystal structure of the complex (blue), the individually crystallized ligand (red), and the dominant conformation from the MD simulation (green). (A) Trypsin/APPI complex (1BRC). (B) HIV-1 NEF/FYN tyrosine kinase SH3 domain complex (1AVZ). (C) Hyhel-5 Fab/lysozyme complex (1BQL). (D) Complex of acetylcholinesterase and fasciculin II (1FSS). (E) Ribonuclease inhibitor/ribonuclease A complex (1DFJ). (F) Subtilisin novo/chymotrypsin inhibitor 2 complex (2SNI), the two most dominant rotamers (in green and magenta), are shown for Ile-56.

Fig. 3.

Latch residues in six complexes. Details are as described for Fig. 2. (A) CheA/CheY complex (1A0O). (B) HIV-1 NEF/FYN tyrosine kinase SH3 domain complex (1AVZ). (C) Subtilisin Carlsberg/Eglin C (1CSE). (D) Acetylcholinesterase/fasciculin II complex (1FSS). (E) Ribonuclease inhibitor/ribonuclease A complex (1DFJ). (F) Subtilisin novo/chymotrypsin inhibitor 2 complex (2SNI). In some cases in which the clarity of the picture is not compromised, the interacting residue on the other side of the interface is also shown as sticks inside the surface representation.

Fig. 4.

Structurally conserved anchor residues for different homologs. Anchor residues are written in red letters. The active site loop from the reference structure is shown with overlap residues from all of the homologs; residues of the same type have the same color. (A) Chymotrypsin inhibitor 2 (from 2SNI) is compared with ligands in PDB ID entries 1LW6, 1SBN, 1MEE, 2SEC, 1ACB, and 3TEC. (B) APPI (from 1BRC) is compared with ligands in PDB ID entries 1BTH, 1BZX, 1CBW, 1EAW, 1F5R, 1FAK, 2KAI, and 1AN1. Here, a small turn in the Lys anchor is observed in the two complexes that inhibit α-chymotrypsin, PDB ID entries 1MTN and 1ACB.