Cyclic nucleotides (cAMP and cGMP) regulate multiple intracellular processes and are thus of a great general interest for molecular and structural biologists. To study the allosteric mechanism of different cyclic nucleotide binding (CNB) domains, we compared cAMP-bound and cAMP-free structures (PKA, Epac, and two ionic channels) using a new bioinformatics method: local spatial pattern alignment. Our analysis highlights four major conserved structural motifs: 1) the phosphate binding cassette (PBC), which binds the cAMP ribose-phosphate, 2) the "hinge," a flexible helix, which contacts the PBC, 3) the beta(2,3) loop, which provides precise positioning of an invariant arginine from the PBC, and 4) a conserved structural element consisting of an N-terminal helix, an eight residue loop and the A-helix (N3A-motif). The PBC and the hinge were included in the previously reported allosteric model, whereas the definition of the beta(2,3) loop and the N3A-motif as conserved elements is novel. The N3A-motif is found in all cis-regulated CNB domains, and we present a model for an allosteric mechanism in these domains. Catabolite gene activator protein (CAP) represents a trans-regulated CNB domain family: it does not contain the N3A-motif, and its long range allosteric interactions are substantially different from the cis-regulated CNB domains.

Here, we introduce a new approach to macromolecular docking which combines the continuous global optimization algorithm CGU [6, 21] with the grid-based conformational search engine DOT [23]. We also detail the use of this method to dock acetylcholine into the fasciculin-acetylcholinesterase complex. Fasciculin blocks access to the long, narrow pathway leading to the active site, which is located in the interior of the acetylcholinesterase. For this reason, finding the location computationally is rather non-trivial. We not only successfully identified the optimal docking configuration, but were able to do so in a matter of hours.

The purpose of the molecular interaction program DOT (Daughter of Turnip) is rapid computation of the electrostatic potential energy between two proteins or other charged molecules. DOT exhaustively tests all six degrees of freedom, rotational and translational, and produces a grid of approximate interaction energies and orientations. It is able to do this because the problem is cast as the convolution of the potential field of the first molecule and any rotated charge distribution of the second. The algorithm lends itself to both parallelization and vectorization, permitting huge increases in computational speed over other methods for obtaining the same information. For example, a complete mapping of interactions between plastocyanin and cytochrome c was done in eight minutes using 256 nodes of an Intel Paragon. DOT is expected to be particularly useful as a rapid screen to find configurations for more detailed study using exact energy models.