R. Veevers, S. Hayward
"Methodological improvements for the analysis of domain movements in large biomolecular complexes" Biophysics and Physicobiology, 16, 328-336, (2019).

Domain movements play a prominent role in the function of many biomolecules such as the ribosome and F0F1-ATP synthase. As more structures of large biomolecules in different functional states become available as experimental techniques for structure determination advance, there is a need to develop methods to understand the conformational changes that occur. DynDom and DynDom3D were developed to analyse two structures of a biomolecule for domain movements. They both used an original method for domain recognition based on clustering of “rotation vectors”. Here we introduce significant improvements in both the methodology and implementation of a tool for the analysis of domain movements in large multimeric biomolecules. The main improvement is in the recognition of domains by using all six degrees of freedom required to describe the movement of a rigid body. This is achieved by way of Chasles’ theorem in which a rigid-body movement can be described as a screw movement about a unique axis. Thus clustering now includes, in addition to rotation vector data, screw-axis location data and axial climb data. This improves both the sensitivity of domain recognition and performance. A further improvement is the recognition and annotation of interdomain bending regions, something not done for multimeric biomolecules in DynDom3D. This is significant as it is these regions that collectively control the domain movement. The new stand-alone, platform-independent implementation, DynDom6D, can analyse biomolecules comprising protein, DNA and RNA, and employs an alignment method to automatically achieve the required equivalence of atoms in the two structures.

C.Girdlestone, S.Hayward
"The DynDom3D webserver for the analysis of domain movements in multimeric proteins" Journal of Computational Biology, 23(1), 21-26, 2016.

DynDom3D is a program for the analysis of domain movements in multimeric proteins. Its inputs are two structure files that indicate a possible domain movement, but the onus has been on the user to process the files so that there is the necessary one-to-one equivalence between atoms in the two atom lists. This is often a prohibitive task to carry out manually, which has limited the application of DynDom3D. Here we report on a webserver with a preprocessor that automatically creates an equivalence between atoms using sequence alignment methods. The processed structure files are passed to DynDom3D and the results are presented on a webpage that includes molecular graphics for easy visualization.

G.Poornam, A. Matsumoto, H.Ishida, S.Hayward
"A method for the analysis of domain movements in large biomolecular complexes" Proteins, 76, 201-21, 2009.

A new method for the analysis of domain movements in large, multichain, biomolecular complexes is presented. The method is applicable to any molecule for which two atomic structures are available that represent a conformational change indicating a possible domain movement. The method is blind to atomic bonding and atom type and can, therefore, be applied to biomolecular complexes containing different constituent molecules such as protein, RNA, or DNA. At the heart of the method is the use of blocks located at grid points spanning the whole molecule. The rotation vector for the rotation of atoms from each block between the two conformations is calculated. Treating components of these vectors as coordinates means that each block is associated with a point in a “rotation space” and that blocks with atoms that rotate together, perhaps as part of the same rigid domain, will have colocated points. Thus a domain can be identified from the clustering of points from blocks that span it. Domain pairs are accepted for analysis of their relative movements in terms of screw axes based upon a set of reasonable criteria. Here, we report on the application of the method to biomolecules covering a considerable size range: hemoglobin, liver alcohol dehydrogenase, S-Adenosylhomocysteine hydrolase, aspartate transcarbamylase, and the 70S ribosome. The results provide a depiction of the conformational change within each molecule that is easily understood, giving a perspective that is expected to lead to new insights. Of particular interest is the allosteric mechanism in some of these molecules. Results indicate that common boundaries between subunits and domains are good regions to focus on as movement in one subunit can be transmitted to another subunit through such interfaces.

S. Hayward, R. A. Lee
"Improvements in the analysis of domain motions in proteins from conformational change: DynDom version 1.50" J Mol Graph Model, Dec, 21(3), 181-3, 2002.

DynDom is a program that analyses conformational change in proteins for dynamic domains, hinge axes, and hinge-bending regions. Here, a number of improvements and additions are reported which have been implemented in the new version 1.50. The most significant improvement is in the determination of the hinge-bending residues. A new routine also compares quantities relating to the main-chain dihedrals of bending residues with the hinge-bending motion.

S. Hayward, H. J. C. Berendsen
"Systematic Analysis of Domain Motions in Proteins from Conformational Change; New Results on Citrate Synthase and T4 Lysozyme" Proteins, Structure, Function and Genetics, 30, 144, 1998.

Methods developed originally to analyze domain motions from simulation [Proteins 27:425–437, 1997] are adapted and extended for the analysis of X-ray conformers and for proteins with more than two domains. The method can be applied as an automatic procedure to any case where more than one conformation is available. The basis of the methodology is that domains can be recognized from the difference in the parameters governing their quasi-rigid body motion, and in particular their rotation vectors. A clustering algorithm is used to determine clusters of rotation vectors corresponding to main-chain segments that form possible dynamic domains. Domains are accepted for further analysis on the basis of a ratio of interdomain to intradomain fluctuation, and Chasles' theorem is used to determine interdomain screw axes. Finally residues involved in the interdomain motion are identified. The methodology is tested on citrate synthase and the M6I mutant of T4 lysozyme. In both cases new aspects to their conformational change are revealed, as are individual residues intimately involved in their dynamics. For citrate synthase the beta sheet is identified to be part of the hinging mechanism. In the case of T4 lysozyme, one of the four transitions in the pathway from the closed to the open conformation, furnished four dynamic domains rather than the expected two. This result indicates that the number of dynamic domains a protein possesses may not be a constant of the motion.

Main DynDom Reference book cover image

S. Hayward, A. Kitao, H. J. C. Berendsen
"Model-Free Methods of Analyzing Domain Motions in Proteins from Simulation: A Comparison of Normal Mode Analysis and Molecular Dynamics Simulation of Lysozyme" Proteins, Structure, Function and Genetics, 27, 425, 1997.

Model-free methods are introduced to determine quantities pertaining to protein domain motions from normal mode analyses and molecular dynamics simulations. For the normal mode analysis, the methods are based on the assumption that in low frequency modes, domain motions can be well approximated by modes of motion external to the domains. To analyze the molecular dynamics trajectory, a principal component analysis tailored specifically to analyze interdomain motions is applied. A method based on the curl of the atomic displacements is described, which yields a sharp discrimination of domains, and which defines a unique interdomain screwaxis. Hinge axes are defined and classified as twist or closure axes depending on their direction. The methods have been tested on lysozyme. A remarkable correspondence was found between the first normal mode axis and the first principal mode axis, with both axes passing within 3 Å of the alpha-carbon atoms of residues 2, 39, and 56 of human lysozyme, and near the interdomain helix. The axes of the first modes are overwhelmingly closure axes. A lesser degree of correspondence is found for the second modes, but in both cases they are more twist axes than closure axes. Both analyses reveal that the interdomain connections allow only these two degrees of freedom, one more than provided by a pure mechanical hinge. Proteins 27:425-437, 1997.

Database References

Daniel Taylor, Gavin Cawley, Steven Hayward
"Quantitative Method for the Assignment of Hinge and Shear Mechanism in Protein Domain Movements" Bioinformatics, 2014.

A popular method for classification of protein domain movements apportions them into two main types: those with a ‘hinge’ mechanism and those with a ‘shear’ mechanism. The intuitive assignment of domain movements to these classes has limited the number of domain movements that can be classified in this way. Furthermore, whether intended or not, the term ‘shear’ is often interpreted to mean a relative translation of the domains. Results:
Numbers of occurrences of four different types of residue contact changes between domains were optimally combined by logistic regression using the training set of domain movements intuitively classified as hinge and shear to produce a predictor for hinge and shear. This predictor was applied to give a 10-fold increase in the number of examples over the number previously available with a high degree of precision. It is shown that overall a relative translation of domains is rare, and that there is no difference between hinge and shear mechanisms in this respect. However, the shear set contains significantly more examples of domains having a relative twisting movement than the hinge set. The angle of rotation is also shown to be a good discriminator between the two mechanisms. Availability and implementation: Results are free to browse at http:// Contact: Supplementary information: Supplementary data are available at Bioinformatics online. Received on January 31, 2014; revised on July 8, 2014; accepted on July 18, 2014

Daniel Taylor, Gavin Cawley, Steven Hayward
"Classification of Domain Movements in Proteins Using Dynamic Contact Graphs" PLOS ONE, 8(11), 2013.

A new method for the classification of domain movements in proteins is described and applied to 1822 pairs of structures from the Protein Data Bank that represent a domain movement in two-domain proteins. The method is based on changes in contacts between residues from the two domains in moving from one conformation to the other. We argue that there are five types of elemental contact changes and that these relate to five model domain movements called: “free”, “open-closed”, “anchored”, “sliding-twist”, and “see-saw.” A directed graph is introduced called the “Dynamic Contact Graph” which represents the contact changes in a domain movement. In many cases a graph, or part of a graph, provides a clear visual metaphor for the movement it represents and is a motif that can be easily recognised. The Dynamic Contact Graphs are often comprised of disconnected subgraphs indicating independent regions which may play different roles in the domain movement. The Dynamic Contact Graph for each domain movement is decomposed into elemental Dynamic Contact Graphs, those that represent elemental contact changes, allowing us to count the number of instances of each type of elemental contact change in the domain movement. This naturally leads to sixteen classes into which the 1822 domain movements are classified.

Guoying Qi, Steven Hayward
"A database of ligand-induced domain movements in enzymes" BMC Structural Biology, 9(13), 2009.

Background: Conformational change induced by the binding of a substrate or coenzyme is a poorly understood stage in the process of enzyme catalysed reactions. For enzymes that exhibit a domain movement, the conformational change can be clearly characterized and therefore the opportunity exists to gain an understanding of the mechanisms involved. The development of the non-redundant database of protein domain movements contains examples of ligand-induced domain movements in enzymes, but this valuable data has remained unexploited. Description: The domain movements in the non-redundant database of protein domain movements are those found by applying the DynDom program to pairs of crystallographic structures contained in Protein Data Bank files. For each pair of structures cross-checking ligands in their Protein Data Bank files with the KEGG-LIGAND database and using methods that search for ligands that contact the enzyme in one conformation but not the other, the non-redundant database of protein domain movements was refined down to a set of 203 enzymes where a domain movement is apparently triggered by the binding of a functional ligand. For these cases, ligand binding information, including hydrogen bonds and salt-bridges between the ligand and specific residues on the enzyme is presented in the context of dynamical information such as the regions that form the dynamic domains, the hinge bending residues, and the hinge axes. Conclusion: The presentation at a single website of data on interactions between a ligand and specific residues on the enzyme alongside data on the movement that these interactions induce, should lead to new insights into the mechanisms of these enzymes in particular, and help in trying to understand the general process of ligand-induced domain closure in enzymes.

Guoying Qi, Richard Lee, Steven Hayward
"A comprehensive and non-redundant database of protein domain movements" Bioinformatics, 21(12):2832-2838, 2005.

Motivation: The current DynDom database of protein domain motions is a user-created database that suffers from selectivity and redundancy. The aim of the analysis presented here was to overcome both these limitations and to produce both a comprehensive and non-redundant description of domain movements from structures stored in the current protein data bank.

Results: A multi-step procedure is applied that starts with grouping proteins in the structural databank into families based on sequence similarity. Multiple sequence alignment, conformational clustering, and a dimensional clustering method based on the Gram-Schmidt algorithm, are applied to members of each family to remove dynamic redundancy in their domain movements. Representative domain movements are described in terms of domains, hinge axes, and hinge-bending residues using the DynDom program. The results show that within an average family of 11.5 members there are on average only 1.31 different domain movements indicating a high redundancy in the movements these structures represent. This verifies earlier findings that domain movements are usually highly controlled. Despite the removal of this considerable redundancy the process has resulted in double the number of domain movements stored in the user-created database. The data is organised in a relational database with a web-interface.

R. A. Lee, M. Razaz, S. Hayward
"The DynDom database of protein domain motions" Bioinformatics, 19, 10, 2003.

A relational database has been developed based on the results from the application of the DynDom program to a number of proteins for which multiple X-ray conformers are available. The database is populated via a web-based tool that allows visitors to the website to run the DynDom program server-side by selecting pairs of X-ray conformers by Protein Data Bank code and chain identifier.


S. Hayward, A. Kitao
"Molecular Dynamics Simulations of NAD+-Induced Domain Closure in Horse Liver Alcohol Dehydrogenase" Biophysical Journal, 91:1823-1831, 2006.

Horse liver alcohol dehydrogenase is a homodimer, the protomer having a coenzyme-binding domain and a catalytic domain. Using all available x-ray structures and 50 ns of molecular dynamics simulations, we investigated the mechanism of NAD+-induced domain closure. When the well-known loop at the domain interface was modeled to its conformation in the closed structure, the NAD+-induced domain closure from the open structure could be simulated with remarkable accuracy. Native interactions in the closed structure between Arg369, Arg47, His51, Ala317, Phe319, and NAD+ were seen to form at different stages during domain closure. Removal of the Arg369 side-chain charge resulted in the loss of the tendency to close, verifying that specific interactions do help drive the domains closed. Further simulations and a careful analysis of x-ray structures suggest that the loop prevents domain closure in the absence of NAD+, and a cooperative mechanism operates between the subunits for domain closure. This cooperative mechanism explains the role of the loop as a block to closure because in the absence of NAD+ it would prevent the occurrence of an unliganded closed subunit when the other subunit closes on NAD+. Simulations that started with one subunit open and one closed supported this.

I. Daidone, D. Roccatano, S. Hayward
"Investigating the accessibility of the closed domain conformation of citrate synthase using essential dynamics sampling" J. Mol. Biol. , 339, 515, 2004.

A molecular dynamics study of pig heart citrate synthase is presented that aims to directly address the question of whether, for this enzyme, the ligand-induced closed domain conformation is accessible to the open unliganded enzyme. The approach utilises the technique of essential dynamics sampling, which is used in two modes. In exploring mode, the enzyme is encouraged to explore domain conformations it might not normally sample in free molecular dynamics simulation. In targeting mode, the enzyme is encouraged to adopt the domain conformation of a target structure. Using both modes extensively, it has been found that when the enzyme is prepared from a crystallographic open-domain structure and is in the unliganded state, it is unable to adopt the crystallographic closed-domain conformation of the liganded enzyme. Likewise, when the enzyme is prepared from the crystallographic closed liganded conformation with the ligands removed, it is unable to adopt the crystallographic open domain conformation. Structural investigations point to a common structural difference that is the source of this energy barrier; namely, the shift of α-helix 328–341 along its own axis relative to the large domain. Without this shift, the domains are unable to close or open fully. The charged substrate, oxaloacetate, binds near the base of this helix in the large domain and the interaction of Arg329 at the base of the helix with oxaloacetate is one that is consistent with the shift of this helix in going from the crystallographic open to closed structure. Therefore, the results suggest that without the substrate the enzyme remains in a partially open conformation ready to receive the substrate. In this way, the efficiency of the enzyme should be increased over one that is closed part of the time, with its binding site inaccessible to the substrate.

S. Hayward
"Identification of specific interactions that drive ligand-induced closure in five enzymes with classic domain movements" J. Mol. Biol. , 339, 1001, 2004.

In order to better understand ligand-induced closure in domain enzymes, open unliganded X-ray structures and closed liganded X-ray structures have been studied in five enzymes: adenylate kinase, aspartate aminotransferase, citrate synthase, liver alcohol dehydrogenase, and the catalytic subunit of cAMP-dependent protein kinase. A sequential model of ligand binding and domain closure was used to test the hypothesis that the ligand actively drives closure from an open conformation. The analysis supports the assumption that each enzyme has a dedicated binding domain to which the ligand binds first and a closing domain. In every case, a small number of residues are identified to interact with the ligand to initiate and drive domain closure. In all cases except adenylate kinase, the backbone of residues located in an interdomain-bending region (hinge site) is identified to interact with the ligand to aid in driving closure. In adenylate kinase, the side-chain of a residue located directly adjacent to a bending region drives closure. It is thought that by binding near a hinge site the ligand is able to get within interaction range of residues when the enzyme is in the open conformation. Interdomain bending regions not involved in inducing closure are involved in control, helping to determine the location of the hinge axis. Similarities have been discovered between aspartate aminotransferase and citrate synthase that only come to light in the context of their dynamical behaviour in response to binding their substrate. Similarity also exists between liver alcohol dehydrogenase and cAMP-dependent protein kinase whereby groups on NAD and ATP, respectively, mimic the backbone of a single amino acid residue in a process where a three residue segment located at the terminus of a β-sheet, moves to form hydrogen bonds with the mimic that resemble those found in a parallel β-sheet. This interaction helps to drive domain closure in a process that has analogy to protein folding.

D. Roccatano, A. E. Mark, S. Hayward
"Investigation of the Mechanism of Domain Closure in Citrate Synthase by Molecular Dynamics Simulation" J. Mol. Biol. , 310, 1039, 2001.

Six, 2 ns molecular dynamics simulations have been performed on the homodimeric enzyme citrate synthase. In three, both monomers were started from the open, unliganded X-ray conformation. In the remaining three, both monomers started from a closed, liganded X-ray conformation, with the ligands removed. Projecting the motion from the simulations onto the experimental domain motion revealed that the free-energy profile is rather flat around the open conformation, with steep sides. The most closed conformations correspond to hinge-bending angles of 12–14 ° compared to the 20 ° that occurs upon the binding of oxaloacetate. It is also found that the open, unliganded X-ray conformation is situated at the edge of the steep rise in free energy, although conformations that are about 5 ° more open were sampled. A rigid-body essential dynamics analysis of the combined open trajectories has shown that domain motions in the direction of the closed X-ray conformation are compatible with the natural domain motion of the unliganded protein, which has just two main degrees of freedom. The simulations starting from the closed conformation suggest a free-energy profile with a small barrier in going from the closed to open conformation. A combined essential dynamics and hinge-bending analysis of a trajectory that spontaneously converts from the closed to open state shows an almost exact correspondence to the experimental transition that occurs upon ligand binding. The simulations support the conclusion from an earlier analysis of the experimental transition that the β-hairpin acts as a mechanical hinge by attaching the small domain to the large domain through a conserved main-chain hydrogen bond and salt-bridges, and allowing rotation to occur via its two flexible termini. The results point to a mechanism of domain closure in citrate synthase that has analogy to the process of closing a door.

S. Hayward
"Structural Principles Governing Domain Motions in Proteins" Proteins, Structure, Function and Genetics, 36, 425, 1999.

With the use of a recently developed method, twenty-four proteins for which two or more X-ray conformers are known have been analyzed to reveal structural principles that govern domain motions in proteins. In all 24 cases, the domain motion is a rotation about a physical axis created through local interactions both covalent and noncovalent. In many cases, two or more mechanical hinges separated in space create a stable hinge axis for precise control of the domain closure. The terminal regions of alpha-helices and beta-sheets have been found to act as mechanical hinges in a significant number of cases. In some cases, the two terminal regions of neighboring strands of a single beta-sheet can create a hinge axis, as can the two termini of a single alpha-helix. These two structures have been termed the "double-hinged beta-sheet" and "double-hinged alpha-helix," respectively. A flexible loop that attaches one domain to another and through which the effective hinge axis passes is another construct that is used to create a hinge. Noncovalent interactions between segments remote along the polypeptide chain can also form hinges. In addition alpha-helices that preserve their hydrogen bonding structure when bent have been found to behave as mechanical hinges. It is suggested that these alpha-helices act as a store of elastic energy that drives the closing of domains for rapid capture of the substrate. If the repertoire of possible interdomain structures is as limited as this study suggests, the dynamic behavior of proteins could soon be predicted using bioinformatics techniques. Proteins 1999;36:425-435.

B. L. de Groot, S. Hayward, D. van Aalten, A. Amadei, H. J. C. Berendsen
"Domain Motions in Bacteriophage T4 Lysozyme; a Comparison between Molecular Dynamics and Crystallographic Data" Proteins, Structure, Function and Genetics, 31, 116, 1998.

A comparison of a series of extended molecular dynamics (MD) simulations of bacteriophage T4 lysozyme in solvent with X-ray data is presented. Essential dynamics analyses were used to derive collective fluctuations from both the simulated trajectories and a distribution of crystallographic conformations. In both cases the main collective fluctuations describe domain motions. The protein consists of an N- and C-terminal domain connected by a long helix. The analysis of the distribution of crystallographic conformations reveals that the N-terminal helix rotates together with either of these two domains. The main domain fluctuation describes a closure mode of the two domains in which the N-terminal helix rotates concertedly with the C-terminal domain, while the domain fluctuation with second largest amplitude corresponds to a twisting mode of the two domains, with the N-terminal helix rotating concertedly with the N-terminal domain. For the closure mode, the difference in hinge-bending angle between the most open and most closed X-ray structure along this mode is 49 degrees. In the MD simulation that shows the largest fluctuation along this mode, a rotation of 45 degrees was observed. Although the twisting mode has much less freedom than the closure mode in the distribution of crystallographic conformations, experimental results suggest that it might be functionally important. Interestingly, the twisting mode is sampled more extensively in all MD simulations than it is in the distribution of X-ray conformations. Proteins 31:116–127,1998