BiGGER: a new (soft) docking algorithm for predicting protein interactions.

A new computationally efficient and automated "soft docking" algorithm is described to assist the prediction of the mode of binding between two proteins, using the three-dimensional structures of the unbound molecules. The method is implemented in a software package called BiGGER (Bimolecular Complex Generation with Global Evaluation and Ranking) and works in two sequential steps: first, the complete 6-dimensional binding spaces of both molecules is systematically searched. A population of candidate protein-protein docked geometries is thus generated and selected on the basis of the geometric complementarity and amino acid pairwise affinities between the two molecular surfaces. Most of the conformational changes observed during protein association are treated in an implicit way and test results are equally satisfactory, regardless of starting from the bound or the unbound forms of known structures of the interacting proteins. In contrast to other methods, the entire molecular surfaces are searched during the simulation, using absolutely no additional information regarding the binding sites. In a second step, an interaction scoring function is used to rank the putative docked structures. The function incorporates interaction terms that are thought to be relevant to the stabilization of protein complexes. These include: geometric complementarity of the surfaces, explicit electrostatic interactions, desolvation energy, and pairwise propensities of the amino acid side chains to contact across the molecular interface. The relative functional contribution of each of these interaction terms to the global scoring function has been empirically adjusted through a neural network optimizer using a learning set of 25 protein-protein complexes of known crystallographic structures. In 22 out of 25 protein-protein complexes tested, near-native docked geometries were found with C(alpha) RMS deviations < or =4.0 A from the experimental structures, of which 14 were found within the 20 top ranking solutions. The program works on widely available personal computers and takes 2 to 8 hours of CPU time to run any of the docking tests herein presented. Finally, the value and limitations of the method for the study of macromolecular interactions, not yet revealed by experimental techniques, are discussed.

Heteronuclear NMR and soft docking: an experimental approach for a structural model of the cytochrome c553-ferredoxin complex.

The combination of docking algorithms with NMR data has been developed extensively for the studies of protein-ligand interactions. However, to extend this development for the studies of protein-protein interactions, the intermolecular NOE constraints, which are needed, are more difficult to access. In the present work, we describe a new approach that combines an ab initio docking calculation and the mapping of an interaction site using chemical shift variation analysis. The cytochrome c553-ferredoxin complex is used as a model of numerous electron-transfer complexes. The 15N-labeling of both molecules has been obtained, and the mapping of the interacting site on each partner, respectively, has been done using HSQC experiments. 1H and 15N chemical shift analysis defines the area of both molecules involved in the recognition interface. Models of the complex were generated by an ab initio docking software, the BiGGER program (bimolecular complex generation with global evaluation and ranking). This program generates a population of protein-protein docked geometries ranked by a scoring function, combining relevant stabilization parameters such as geometric complementarity surfaces, electrostatic interactions, desolvation energy, and pairwise affinities of amino acid side chains. We have implemented a new module that includes experimental input (here, NMR mapping of the interacting site) as a filter to select the accurate models. Final structures were energy minimized using the X-PLOR software and then analyzed. The best solution has an interface area (1037.4 A2) falling close to the range of generally observed recognition interfaces, with a distance of 10.0 A between the redox centers.

The structure of an electron transfer complex containing a cytochrome c and a peroxidase.

Efficient biological electron transfer may require a fluid association of redox partners. Two noncrystallographic methods (a new molecular docking program and 1H NMR spectroscopy) have been used to study the electron transfer complex formed between the cytochrome c peroxidase (CCP) of Paracoccus denitrificans and cytochromes c. For the natural redox partner, cytochrome c550, the results are consistent with a complex in which the heme of a single cytochrome lies above the exposed electron-transferring heme of the peroxidase. In contrast, two molecules of the nonphysiological but kinetically competent horse cytochrome bind between the two hemes of the peroxidase. These dramatically different patterns are consistent with a redox active surface on the peroxidase that may accommodate more than one cytochrome and allow lateral mobility.

The surface-charge asymmetry and dimerisation of cytochrome c550 from Paracoccus denitrificans--implications for the interaction with cytochrome c peroxidase.

The implications of the dimeric state of cytochrome c550 for its binding to Paracoccus cytochrome c peroxidase and its delivery of the two electrons required to restore the active enzyme during catalysis have been investigated. The amino acid sequence of cytochrome c550 of Paracoccus denitrificans strain LMD 52.44 was determined and showed 21 differences from that of strain LMD 22.21. Based on the X-ray structure of the latter, a structure for the cytochrome c550 monomer from strain 52.44 is proposed and a dipole moment of 945 debye was calculated with an orientation close to the exposed haem edge. The behaviour of the cytochrome on molecular-exclusion chromatography is indicative of an ionic strength-dependent monomer (15 kDa)/dimer (30 kDa) equilibrium that can also be detected by 1H-NMR spectroscopy. The apparent mass of 50 kDa observed at very low ionic strength was consistent with the presence of a strongly asymmetric dimer. This was confirmed by cross-linking studies, which showed that a cross-linked species of mass 30 kDa on SDS behaved with an apparent mass of 50 kDa on molecular-exclusion chromatography. A programme which carried out and evaluated molecular docking of two monomers to give a dimer generated a most probable dimer in which the monomer dipoles lay almost antiparallel to each other. The resultant dipole moment of the dimer is therefore small. Although this finding calls into question the possibility of preorientation of a strongly asymmetrically charged cytochrome as it collides with a redox partner, the stoichiometry of complex formation with cytochrome c peroxidase as studied by 1H-NMR spectroscopy shows that it is the monomer that binds.