Community-wide blind prediction experiments such as CAPRI and CASP provide an objective measure of the current state of predictive methodology. Here we describe a community-wide assessment of methods to predict the effects of mutations on protein-protein interactions. Twenty-two groups predicted the effects of comprehensive saturation mutagenesis for two designed influenza hemagglutinin binders and the results were compared with experimental yeast display enrichment data obtained using deep sequencing. The most successful methods explicitly considered the effects of mutation on monomer stability in addition to binding affinity, carried out explicit side chain sampling and backbone relaxation, and evaluated packing, electrostatic and solvation effects, and correctly identified around a third of the beneficial mutations. Much room for improvement remains for even the best techniques, and large-scale fitness landscapes should continue to provide an excellent test bed for continued evaluation of methodological improvement.

To uncover the adaptation mechanisms of photosystems for halotolerance, changes in stoichiometry and activity of photosystems in response to changes of salinities were examined in a halotolerant cyanobacterium, Aphanothece halophytica. Photosynthetic O2 evolution was high even at high salinities. O2 evolution activity increased with increasing external concentration of NaCl, reached a maximum at 1.5 M NaCl, and then decreased. Similar salt dependence was observed for photosystem II activity. On the other hand, photosystem I activity increased concomitantly with increase in salinity. Photoacoustic measurements indicated that appreciable energy storage by photosystem I mediated cyclic electron flow at high salinities. Significant electron donation to photosystem I reaction centres through NAD(P)H-dehydrogenase complexes was observed in high salt media. The contents of cytochrome b6/f and photosystem II were almost constant under various salinity conditions, whereas the levels of chlorophyll α, photosystem I, soluble cytochrome c-553, and NAD(P)H-dehydrogenase increased in the cells grown with high salinities. These results indicate that salt specifically induces an increase of protein levels involving cyclic electron flow around photosystem I that may entail an important role for adaptation of Aphanothece halophytica cells to high salinities.

Precise and rapid calculation of long-range interactions is of crucial importance for molecular dynamics (MD) and Monte Carlo simulations. Instead of the Ewald method or its high speed variant, PME, we applied our novel method, called the force-switching Wolf method, to computation of the free energy landscapes of a short peptide in explicit water. Wolf and co-workers showed that long-range electrostatic energy under a periodic boundary condition can be well reproduced even by truncating the contribution from the distant charges, when the charge neutrality is taken into account. We recently applied the procedure proposed by Wolf and co-workers to a mathematically consistent MD theory by means of a force-switching scheme, and we show that the total electrostatic energy for sodium chloride liquid was well conserved and stable during the MD simulation with the force-switching Wolf method. Our current results for an aqueous peptide solution with a series of canonical and multicanonical molecular dynamics simulations show that the force-switching Wolf method is not only in good accordance with the energies and forces calculated by the conventional PME method but also properly reproduces the solvation and the free energy landscapes of the peptide at 300 K.

Calmodulin (CaM) is a Ca2+-binding messenger protein having four Ca2+-binding motifs named 'EF-hand'; the EF-hand motifs undergo a conformation change induced by Ca2+-binding. In order to study how Ca2+-binding induces the conformation change of EF-hand motifs and which residues are involved in the reaction, two 1μ second long MD simulations were independently performed from the apo- and holo-CaM and their structures and interactions were compared. The Ca2+-binding weakens the helix-helix interaction in all EF-hand, however, the holo-CaM MD adopted the close-like form. The correlation coefficients obtained from the two MDs show the residues comprising interactions being involved in their close-open conformation changes; most of these residues are hydrophobic amino acids but some of them are hydrophilic (T34, H107, N111 and Q143). The hydrophilic residues are expected to lock the EF-hands by their side-chains and main-chain carbonyl oxygen of another hydrophobic residue. Furthermore, the interaction pattern of EF-hand3 and 4 are similar to each other. On the other hand, the interaction pattern of EF-hand2 is different from others; its polar residues are expected to play an important role in regulating the EF-hand2 conformation.

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