Reliable determination of the quaternary structure of a protein is often crucial to a full understanding of its function. However, for decades, crystallographers have sometimes struggled to distinguish between biologically meaningful interfaces observed in a crystal structure, and the unnatural lattice contacts that allow for crystal formation. In order to solve this problem, Eugene Krissinel developed PISA, and later jsPISA, a CCP4 tool that sorts different candidate quaternary structures according to a likelihood obtained from results on dissociation free energy [1, 2]. Additionally, jsPISA incorporates an "interaction radar" that allows for a rapid visualisation of how likely an interface is according to a number of physico-chemical parameters. In order to provide jsPISA with an additional -- independent -- source of information to determine the probability of a given interface to biologically exist, we propose the use of evolutionary covariance data. This proposal is based upon the observation that pairs of residues whose interaction contributes to a biologically important interface are constrained in their evolution [3, 4]. Consequently, the detection of a pair covariation signal points at the existence of a contact between the two residues contributing to the formation of a biologically important interface. The new extension, named PISACov, aims to enhance the results currently displayed by jsPISA with an additional score and new data based on evolutionary covariance analysis, thereby helping determine the relevant quaternary structure in difficult cases.

Structure solution in macromolecular crystallography is not always a straightforward process and it may be rather difficult for structural biologists without advanced training.A trained crystallographer exploits an extended set of approaches and tricks, based on the analysis of several indicators, general assessment of the case, and developed strategies for dealing with a particular class of problems.If such an approach, used by an expert, can be formalised in terms of an algorithm, then it can be implemented as a computer program or automated user advice system to help users solving structures quicker and with a higher success rate.It is not surprising then, that programs for macromolecular crystallography are moving towards full automation, taking off burden from researchers and lowering the entry barriers for novice users.During the last few years, substantial progress has been made towards automation of the whole macromolecular structure determination process.There are a number of examples of successful automatic solutions for various stages of structure determination, such as molecular replacement, experimental phasing, and refinement [1][2][3][4][5].In this communication, we report two novel automation features implemented in CCP4 Cloud [6], the new system for solving macromolecular structures online, released with CCP4 Software Suite 7.1 in 2020.Automated user advice framework, named Verdicts, provides simple graphical representation of results quality with detailed analysis of points for improvement as part of every task (e.g., refinement) report.The analysis includes suggestions on what could be done in order to improve the result (i.e., which parameters could be optimised).Then, the task can be re-run with the suggested parameters, which can be further adjusted by the user as appropriate.Another automation feature, Workflows, was designed for unfolding structure solution Projects, or their parts, automatically using user-supplied data.Such automatically initiated and unfolded Projects may include a number of tasks, arranged in branching Project Trees as if this were done by the user themselves.In common cases without complications, this may result in structure solved, and if not, then a starting Project is offered to the user for analysis and further manipulations, where simple, first-order structure solution attempts are already performed.Any task or branch of the starting Project may be cloned and re-run with optimized parameters, and new tasks may be added as needed.Workflows combine automation and human expert skills, and, therefore, represent an excellent starting point for users with different level of expertise, ranging from novices to experienced crystallographers.Workflows are particularly useful in a common case of processing large sets of isomorphous crystals, because, once structure is solved in one crystal, the process is well-repeatable in systems with moderate modifications.

"In 2013 MX beamlines at the Diamond synchrotron deployed an automated software pipeline, called DIMPLE, for rapid processing of crystals that contain a known protein and possibly a ligand bound. DIMPLE takes the already known ""apo"" structure for the target protein, compares it with the electron density map from X-ray diffraction images, and visualizes areas of the electron density unaccounted for by the structure model. When processing batches of crystals, such feedback allows the user to better decide what to measure next which leads to a more efficient use of the beam time. This year we've enhanced the pipeline to cover more complex cases, including changes in the space group and some changes in conformation. With multiple molecular replacement computations run in parallel, the time from shooting to viewing the difference map is still only a few minutes. While the software is developed primarily for use at synchrotron beamlines, it is included in the CCP4 suite and can be used as well for in-house automation."

The new CCP4 Coordinate Library is a development aiming to provide a common layer of coordinate-related functionality to the existing applications in the CCP4 suite, as well as a variety of tools that can simplify the design of new applications where they relate to atomic coordinates. The Library comprises a wide spectrum of useful functions, ranging from parsing coordinate formats and elementary editing operations on the coordinate hierarchy of biomolecules, to high-level functionality such as calculation of secondary structure, interatomic bonds, atomic contacts, symmetry transformations, structure superposition and many others. Most of the functions are available in a C++ object interface; however, a Fortran interface is provided for compatibility with older CCP4 applications. The paper describes the general principles of the Library design and the most important functionality. The Library, together with documentation, is available under the LGPL license from the CCP4 suite version 5.0 and higher.

The present paper describes the SSM algorithm of protein structure comparison in three dimensions, which includes an original procedure of matching graphs built on the protein's secondary-structure elements, followed by an iterative three-dimensional alignment of protein backbone Calpha atoms. The SSM results are compared with those obtained from other protein comparison servers, and the advantages and disadvantages of different scores that are used for structure recognition are discussed. A new score, balancing the r.m.s.d. and alignment length Nalign, is proposed. It is found that different servers agree reasonably well on the new score, while showing considerable differences in r.m.s.d. and Nalign.

The E-MSD macromolecular structure relational database (http://www.ebi.ac.uk/msd) is designed to be a single access point for protein and nucleic acid structures and related information. The database is derived from Protein Data Bank (PDB) entries. Relational database technologies are used in a comprehensive cleaning procedure to ensure data uniformity across the whole archive. The search database contains an extensive set of derived properties, goodness-of-fit indicators, and links to other EBI databases including InterPro, GO, and SWISS-PROT, together with links to SCOP, CATH, PFAM and PROSITE. A generic search interface is available, coupled with a fast secondary structure domain search tool.

In this work, we present an experimental study of the energy dependence of the photolysis of aqueous HOCl and the subsequent recombination of the OH and Cl photofragments. Using femtosecond transient absorption spectroscopy, we follow the time-dependent concentration of the fragments. The excess energy in the photolysis is given to the fragments as kinetic energy, and tuning the wavelength enables a study of the energy-dependent thermalization and geminate recombination of the fragments in the liquid environment. The recombination yield and rate are governed by the initial fragment separation, and we deduce an approximately linear dependence of the fragment separation versus the kinetic energy with a slope of 0.13 nm/eV. Performing a simple MD simulation of the system gives qualitative agreement with the observations, although the calculated slope is only 0.04 nm/eV.

A theoretical analysis is presented of the problem of how distance-dependent electron transfer in photoinduced forward electron transfer followed by geminate backward electron transfer in liquid solution is re ̄ ected in the viscosity dependence of the magnetic ® eld e€ ect (MFE) on the e ciency of free radical formation u ce in such reactions. The stochastic Liouville equation formalism is employed to model the reaction behaviour of distance-distributed, triplet-born radical pairs (RPs) undergoing free di€ usion, distanceand spin-dependent backward electron transfer, coherent and incoherent spin evolution in the ps time domain. In comparison with real systems the spin situation is simpli® ed by reducing it to a two state S , T 0 problem, yet it is parametrized in a way that allows sensible comparison of the results with those of recent experiments. It is predicted that the MFE on u ce exhibits characteristic minima in the MFE versus viscosity curves, and it is veri® ed in detail that this feature is peculiar to the di€ usional model with distance-dependent electron transfer, i.e. cannot be reproduced with the simpler (`exponential’ ) RP model employing distance-independent rate constants. Thus, the MFE versus viscosity curves are established as a genuine ® ngerprint of distance-dependent electron transfer. The theoretical results compare favourably with recent experimental results obtained with Ru complex/methylviologen RPs.

The magnetic-field dependence of the cage escape efficiency (phi(ce)) of [Ru(bpy)(3)](3+) and methyl viologen radicals (MV+.) from the primary redox pair formed upon quenching of photoexcited [Ru(bpy)(3)](2+) by MV2+ was measured by laser flash spectroscopy in aqueous solution as a function of the magnetic field (0-2.85 T) in the temperature range from 5 to 69 degreesC. Furthermore, the H-1 NMR T-1 times of the paramagnetic [Ru(bpy)(3)](3+) were measured between -40 and 42 degreesC. The kinetic data were analyzed in terms of a kinetic model that takes into account spin conservation in the forward reaction between the (MLCT)-M-3 state of [Ru(bpy)(3)](2+) and the electron acceptor MV2+ yielding a triplet spin-correlated radical pair (RP) and the in-cage backward electron transfer requiring singlet character of the RP. The triplet-to-singlet spin conversion of the geminate RP is explicitly treated by the stochastic Liouville equation formalism. By theoretical simulation of the observed magnetic field dependence of phi(ce), the temperature dependent absolute values of the rate constants k(ce) (cage escape), k(bet) (backward electron transfer in singlet RPs), and k(TS) (magnetic-field independent triplet-to-singlet interconversion) could be assessed. The temperature dependence of k(ce) exhibits a very good proportionality to the solvent viscosity. The values obtained for k(TS) are in good agreement with the results on the electron spin relaxation time of [Ru(bpy)(3)](3+) derived by the Solomon relation from the H-1 NMR T-1 times. The effective rate of backward electron transfer in the geminate RP turns out to be close to spin-controlled, i.e., it is determined by the rate constant k(TS) of the triplet-singlet spin conversion process. The true rate constant k(bet), varying from 5.5 x 10(10) s(-1) to 1.2 x 10(11) s(-1), is about seven times larger than the effective value for the total backward electron transfer comprising spin conversion and spin-allowed backward electron transfer.

High throughput structural genomics projects are now underway.These projects will collect comprehensive data on protein structure.The e-msd has contributed to the detailed data representation model(s) and exchange data formats and mechanisms between each step in a structure determination.This model, incorporates the means for making the information reliable, accurate and up to date and to include indicators of reliability.The e-msd recognizes that in order for data to be used efficiently for searches within the database, and have ensured data uniformity in that the meta-description of the data is consistent across all entries.The e-msd not only allows for data harvesting and archival of clean cross referenced structural data, search mechanisms are being developed to allow research workers to for example automatically annotate structure motifs and binding site properties.

The structures of NinAlp clusters of all compositions with n+p=12, 13, and 14 are studied both experimentally and theoretically. Experimental reactions of the clusters with N2 are used to determine the number of Ni atoms residing in the cluster surfaces. In agreement with the theoretical predictions, the N2 saturation levels are consistent with clusters having icosahedral and icosahedral-based structures. The various N2 adsorption channels seen in the experiment are explained in terms of the computed composition-dependent patterns of the configurational energies of the different structural forms.

The bimolecular ionization of photoexcited molecules is theoretically investigated assuming the light pumping of moderate intensity is either instantaneous or permanent. The kinetics of energy quenching and ion-radical accumulation and recombination after delta -pulse excitation are studied beyond the rate concept, in the framework of Integral Encounter Theory (IET). The results are compared with those obtained within extended Unified Theory (UT), contact and Markovian approximations, and a widely accepted exponential model. When there is a shortage of accepters the theory becomes nonlinear and discloses the striking effect of electron-transfer saturation. In such conditions and under permanent illumination IET is the sole formalism appropriate for a full time-scale (non-Markovian) description of system relaxation. The original program for solving nonlinear IET equations for particle concentrations was developed and first used to calculate the kinetics of relaxation to equilibrium and to a stationary regime. The non-Markovian corrections to the quantum yields of fluorescence and charge separation obtained numerically are in good correspondence with analytic estimates of these quantities.

Kinetic analyses of geminate radical escape yields in terms of a simple (“exponential”) reaction scheme with first-order rate constants of separation and geminate recombination have been widely used in the literature, e.g. to evaluate rate constants of reverse electron transfer (k−et). Here we demonstrate the limited value of such rate constants by formally analysing, in terms of the exponential model, the diffusion coefficient (iz. viscosity) dependence of the radical escape yield as theoretically calculated in the framework of diffusion-dependent electron transfer theory (unified treatment of non-contact photoinduced forward and geminate reverse electron transfer). It is shown that, while the true electron transfer rate constant is kept constant, the apparent rate constant k−et from the exponential model undergoes a wide variation as a function of diffusion coefficient and the rate of spin conversion. Nevertheless, the function k−et(D) represented in a double log plot for various rates of spin conversion provides a useful map suitable to assign characteristic regions of diffusional, spin and reaction control of the geminate process. As an application to real systems the experimental example of the [Ru(bpy)3]2+/methylviologen system is reconsidered. Here a magnetic field effect on the k−et(D) dependence is useful to corroborate the non-contact formation of the radical pair in the photochemical forward electron transfer reaction.