Molecular-dynamics simulations of [Met5]- and [D-Ala2,Met5]-enkephalins. Biological implication of monomeric folded and dimeric unfolded conformations.

To investigate the biologically active conformation of enkephalin, molecular-dynamics simulations were applied to [Met5]- and [D-Ala2,Met5]-enkephalins. The dynamic trajectory of monomeric extended [Met5]-enkephalin was analysed in terms of relative mobility between respective torsions of backbone chain. After 10 ps of the dynamics simulation, the conformational transition was converged into a stationary state among the beta-bend folded forms, where they are stabilized by several intramolecular hydrogen-bond formations. Similar conformational transition was also observed in the dynamics simulation of [D-Ala2,Met5]enkephalin, which is a more mu-receptor-specific peptide than [Met5]enkephalin. The geometrical correspondence between the monomeric enkephalin conformation in the stationary state and morphine molecule (a mu-specific rigid opiate) was surveyed by virtue of the triangular substructures generated by choosing three functional atoms in each molecule, and good resemblances were observed. On the other hand, the dynamics simulation of the antiparallel extended [Met5]enkephalin dimer showed a trajectory different from that of the monomeric one. Two intermolecular hydrogen bonds at Tyr1 (NH3+)...Met5(CO2-) end residues were held throughout the 100 ps simulation, the dimeric structure being consequently kept. The conformational transition of the backbone chains from the antiparallel extended form to the twisted one took place via an intermediate state. Many conformations revealed during the dynamics simulation showed that the relative orientations of each two Tyr1, Gly3, Phe4 and Met5 residues in the dimer are nearly related by a pseudo-C2-symmetry respectively, and both halves of the dimer structure could be further fitted to the monomeric folded enkephalin conformation. The monomeric and dimeric conformations of enkephalin at their stationary states are discussed in relation to the substrate-specificity for mu- and delta-opioid receptors.     

Two gamma-bends in the backbone conformation of [D-Ala2]-leucine enkephalin in solution

The solution conformation of [D-Ala2]-leucine enkephalin in its zwitterionic form in DMSO-d6 has been monitored by one- and two-dimensional proton magnetic resonance spectroscopy at 500 MHz. The resonances from the labile amide protons and the nonlabile protons have been assigned from the shift correlated spectroscopy. The chemical shift of the amide and C-alpha protons are found to vary with temperature but in opposite directions, except the C-alpha proton of the terminal tyrosine residue. This behavior has been explained by the shifting of equilibrium between the zwitterionic and neutral forms of the [D-Ala2]-leucine enkephalin and probably conformational changes accompanying temperature variation. The low values of the temperature coefficients of leucine and glycine amide protons indicate that these protons are either intramolecularly hydrogen bonded or solvent shielded. The observation of sequential cross peaks in the nuclear Overhauser effect spectra obtained at various mixing times, tau m (200-900 ms), indicate an extended backbone, which does not corroborate with the presence of a folded structure, i.e., beta-bend type structure. The estimate of interproton distances in conjunction with the low values of temperature coefficients of the leucine and glycine amide protons and vicinal coupling constants 3JHN-C alpha H have been rationalized by the predominance of two gamma-bends in the backbone conformation of [D-Ala2]-leucine enkephalin. The gamma-bend around the D-Ala residue has phi = 80 degrees and psi = 270 degrees, while the one around Phe it has phi = 285 degrees and psi = 90 degrees.     

1H NMR assignments of apo calcyclin and comparative structural analysis with calbindin D9k and S100 beta.

The homodimeric S100 protein calcyclin has been studied in the apo state by two-dimensional 1H NMR spectroscopy. Using a combination of scalar correlation and NOE experiments, sequence-specific 1H NMR assignments were obtained for all but one backbone and > 90% of the side-chain resonances. To our knowledge, the 2 x 90 residue (20 kDa) calcyclin dimer is the largest protein system for which such complete assignments have been made by purely homonuclear methods. Sequential and medium-range NOEs and slowly exchanging backbone amide protons identified directly the four helices and the short antiparallel beta-type interaction between the two binding loops that comprise each subunit of the dimer. Further analysis of NOEs enabled the unambiguous assignment of 556 intrasubunit distance constraints, 24 intrasubunit hydrogen bonding constraints, and 2 x 26 intersubunit distance constraints. The conformation of the monomer subunit was refined by distance geometry and restrained molecular dynamics calculations using the intrasubunit constraints only. Calculation of the dimer structure starting from this conformational ensemble has been reported elsewhere. The extent of structural homology among the apo calcyclin subunit, the monomer subunit of apo S100 beta, and monomeric apo calbindin D9k has been examined in detail by comparing 1H NMR chemical shifts and secondary structures. This analysis was extended to a comprehensive comparison of the three-dimensional structures of the calcyclin monomer subunit and calbindin D9k, which revealed greater similarity in the packing of their hydrophobic cores than was anticipated previously. Together, these results support the hypothesis that all members of the S100 family have similar core structures and similar modes of dimerization. Analysis of the amphiphilicity of Helix IV is used to explain why calbindin D9k is monomeric, but full-length S100 proteins form homodimers.     

Three-dimensional solution structure of an insulin dimer. A study of the B9(Asp) mutant of human insulin using nuclear magnetic resonance, distance geometry and restrained molecular dynamics.

The solution structure of the B9(Asp) mutant of human insulin has been determined by two-dimensional 1H nuclear magnetic resonance spectroscopy. Thirty structures were calculated by distance geometry from 451 interproton distance restraints based on intra-residue, sequential and long-range nuclear Overhauser enhancement data, 17 restraints on phi torsional angles obtained from 3JH alpha HN coupling constants, and the restraints from 17 hydrogen bonds, and the three disulphide bridges. The distance geometry structures were optimized using restrained molecular dynamics (RMD) and energy minimization. The average root-mean-square deviation for the best 20 RMD refined structures is 2.26 A for the backbone and 3.14 A for all atoms if the less well-defined N and C-terminal residues are excluded. The helical regions are better defined, with root-mean-square deviation values of 1.11 A for the backbone and 2.03 A for all atoms. The data analysis and the calculations show that B9(Asp) insulin, in water solution at the applied pH (1.8 to 1.9), is a well-defined dimer with no detectable difference between the two monomers. The association of the two monomers in the solution dimer is relatively loose as compared with the crystal dimer. The overall secondary and tertiary structures of the monomers in the 2Zn crystal hexamer is found to be preserved. The conformation-averaged NMR structures obtained for the monomer is close to the structure of molecule 1 in the hexamer of the 2Zn insulin crystal. However, minor, but significant deviations from this structure, as well as from the structure of monomeric insulin in solution, exist and are ascribed to the absence of the hexamer and crystal packing forces, and to the presence of monomer-monomer interactions, respectively. Thus, the monomer in the solution dimer shows a conformation similar to that of the crystal monomer in molecular regions close to the monomer-monomer interface, whereas it assumes a conformation similar to that of the solution structure of monomeric insulin in other regions, suggesting that B9(Asp) insulin adopts a monomer-like conformation when this is not inconsistent with the monomer-monomer arrangement in the dimer.     

MD studies of the DIS/DIS kissing complex solution and x-ray structures

As in all retroviruses, human immunodeficiency virus (HIV) genomic RNA is packaged into virions as a dimer. The two copies of the genome are noncovalently linked by their 5'-ends in the dimerization initiating site (DIS), which folds as a hairpin containing an apical autocomplementary sequence. In vitro, DIS is able to dimerize in two conformations: a kissing complex and an extended dimer. Both conformations have been resolved by NMR and x-ray diffraction. Here, we report molecular dynamics (MD) studies of the two available structures for the DIS/DIS kissing complex in aqueous solution and in the presence of sodium counterions. The two structures behave in two different manners. On one hand, the NMR structure displays a very stable behavior, and the simulated structure remains very close to the starting structure. On the other hand, the structure issued from crystallography displays a more dynamic behavior, in which residues A8 and A9 are seen in a new and surprising bulge-in conformation. The transition from the bulge-out to the bulge-in conformation is analyzed, and a new and simple dimerization process is proposed.     

The N-Terminal Actin-Binding Tandem Calponin-Homology (CH) Domain of?Dystrophin Is in a Closed Conformation in Solution and When Bound to?F-actin

Deficiency of the vital muscle protein dystrophin triggers Duchenne/Becker muscular dystrophy, but the structure-function relationship of dystrophin is poorly understood. To date, molecular structures of three dystrophin domains have been determined, of which the N-terminal actin-binding domain (N-ABD or ABD1) is of particular interest. This domain is composed of two calponin-homology (CH) domains, which form an important class of ABDs in muscle proteins. A previously determined x-ray structure indicates that the dystrophin N-ABD is a domain-swapped dimer, with each monomer adopting an extended, open conformation in which the two CH domains do not interact. This structure is controversial because it contradicts functional studies and known structures of similar ABDs from other muscle proteins. Here, we investigated the solution conformation of the dystrophin N-ABD using a very simple and elegant technique of pyrene excimer fluorescence. Using the wild-type protein, which contains two cysteines, and the corresponding single-cysteine mutants, we show that the protein is a monomer in solution and is in a closed conformation in which the two CH domains seem to interact, as observed from the excimer fluorescence of pyrene-labeled wild-type protein. Excimer fluorescence was also observed in its actin-bound form, indicating that the dystrophin N-ABD binds to F-actin in a closed conformation. Comparison of the dystrophin N-ABD conformation with other ABDs indicates that the tandem CH domains in general may be monomeric in solution and predominantly occur in closed conformation, whereas their actin-bound conformations may differ.     

Small-angle X-ray Scattering of Apolipoprotein A-IV Reveals the Importance of Its Termini for Structural Stability

ApoA-IV is an amphipathic protein that can emulsify lipids and has been linked to protective roles against cardiovascular disease and obesity. We previously reported an x-ray crystal structure of apoA-IV that was truncated at its N and C termini. Here, we have extended this work by demonstrating that self-associated states of apoA-IV are stable and can be structurally studied using small-angle x-ray scattering. Both the full-length monomeric and dimeric forms of apoA-IV were examined, with the dimer showing an elongated rod core with two nodes at opposing ends. The monomer is roughly half the length of the dimer with a single node. Small-angle x-ray scattering visualization of several deletion mutants revealed that removal of both termini can have substantial conformational effects throughout the molecule. Additionally, the F334A point mutation, which we previously showed increases apoA-IV lipid binding, also exhibited large conformational effects on the entire dimer. Merging this study''s low-resolution structural information with the crystal structure provides insight on the conformation of apoA-IV as a monomer and as a dimer and further defines that a clasp mechanism may control lipid binding and, ultimately, protein function.     

Studies of peptides forming 3(10)- and alpha-helices and beta-bend ribbon structures in organic solution and in model biomembranes by Fourier transform infrared spectroscopy

In order to examine the potential correlation between infrared absorption spectra and 3(10)- and alpha-helices and beta-bend ribbon structures, the secondary structures of synthetic peptides known to contain pure 3(10)-helices, mixed 3(10)/alpha-helices, and pure beta-bend ribbon structures, based upon X-ray diffraction and NMR studies, have been investigated by using FTIR spectroscopy incorporating resolution-enhancement techniques. Studies of the peptides known to contain a stable 3(10)-helix in CDCl3 show the main amide I band of fully stable 3(10)-helices occurs at 1666-1662 cm-1. Resolution-enhancement methods revealed small contributions at 1681-1678 and 1646-1644 cm-1, while the amide II band occurs at 1533-1531 cm-1. Peptides known to contain both alpha- and 3(10)-helices in their structure exhibit bands characteristic of both types of conformation. Peptides known to fold into the beta-bend ribbon structure show an amide I band maximum at 1648-1645 cm-1 with the amide II band at 1538-1536 cm-1. Incorporation of these peptides into model membrane structures, e.g., DMPC vesicles, in aqueous buffer sometimes produces changes in the peptide secondary structure. Those peptides which possess a 3(10)-helical structure in CDCl3 solution change the secondary structure in DMPC vesicles to predominantly alpha-helical, plus a contribution from short, unstable 3(10)-helix and/or beta-turns. Those peptides which contain a combination of alpha- and 3(10)-helical structures in CDCl3 solution tend to retain some 3(10)-helical structure within the lipid environment, although the overall H-bonding pattern is altered. Those peptides which form a beta-bend ribbon structure appear to be largely unaffected in the membrane environment.(ABSTRACT TRUNCATED AT 250 WORDS)     

Molecular dynamics simulation of dimeric and monomeric forms of human prion protein: insight into dynamics and properties

A central theme in prion protein research is the detection of the process that underlies the conformational transition from the normal cellular prion form (PrP(C)) to its pathogenic isoform (PrP(Sc)). Although the three-dimensional structures of monomeric and dimeric human prion protein (HuPrP) have been revealed by NMR spectroscopy and x-ray crystallography, the process underlying the conformational change from PrP(C) to PrP(Sc) and the dynamics and functions of PrP(C) remain unknown. The dimeric form is thought to play an important role in the conformational transition. In this study, we performed molecular dynamics (MD) simulations on monomeric and dimeric HuPrP at 300 K and 500 K for 10 ns to investigate the differences in the properties of the monomer and the dimer from the perspective of dynamic and structural behaviors. Simulations were also undertaken with Asp178Asn and acidic pH, which is known as a disease-associated factor. Our results indicate that the dynamics of the dimer and monomer were similar (e.g., denaturation of helices and elongation of the beta-sheet). However, additional secondary structure elements formed in the dimer might result in showing the differences in dynamics and properties between the monomer and dimer (e.g., the greater retention of dimeric than monomeric tertiary structure).     

Molecular dynamics simulations of the amyloid-beta binding alcohol dehydrogenase (ABAD) enzyme

In this work, we present 10 ns molecular dynamics simulations of the homotetramer of the ABAD enzyme, as well as of the structural units, dimer and monomer, that assemble to form the tetramer, in the presence and absence of a NAD-inhibitor adduct. The aim was to compare the stability of the different structures and to study the effects of the inhibitor binding on the flexibility of the enzyme structure. The results indicate that the tetramer, dimer and monomer show a comparable stability and that tetramerization stabilizes some regions of the protein that when exposed to the solvent in dimer and monomer become more flexible. Binding of the cofactor and inhibitor stabilizes the protein, the main effect being a stabilization of the substrate binding loop. In the absence of the ligand, this region was found to have a much higher flexibility and to adopt an open conformation. An interesting result emerging from this work is the conformational flexibility exhibited by the azepane and benzene rings of the inhibitor moiety of the adduct, which appears to be influenced by the mobility of the substrate binding loop. This highlights the importance of integrate the flexibility of the substrate binding loop into de novo design of inhibitors of ABAD.     

Computational study of the fibril organization of polyglutamine repeats reveals a common motif identified in beta-helices

The formation of fibril aggregates by long polyglutamine sequences is assumed to play a major role in neurodegenerative diseases such as Huntington. Here, we model peptides rich in glutamine, through a series of molecular dynamics simulations. Starting from a rigid nanotube-like conformation, we have obtained a new conformational template that shares structural features of a tubular helix and of a beta-helix conformational organization. Our new model can be described as a super-helical arrangement of flat beta-sheet segments linked by planar turns or bends. Interestingly, our comprehensive analysis of the Protein Data Bank reveals that this is a common motif in beta-helices (termed beta-bend), although it has not been identified so far. The motif is based on the alternation of beta-sheet and helical conformation as the protein sequence is followed from the N to the C termini (beta-alpha(R)-beta-polyPro-beta). We further identify this motif in the ssNMR structure of the protofibril of the amyloidogenic peptide Abeta(1-40). The recurrence of the beta-bend suggests a general mode of connecting long parallel beta-sheet segments that would allow the growth of partially ordered fibril structures. The design allows the peptide backbone to change direction with a minimal loss of main chain hydrogen bonds. The identification of a coherent organization beyond that of the beta-sheet segments in different folds rich in parallel beta-sheets suggests a higher degree of ordered structure in protein fibrils, in agreement with their low solubility and dense molecular packing.     

Insight into the activity of SARS main protease: Molecular dynamics study of dimeric and monomeric form of enzyme

The phenomenon that SARS coronavirus main protease (SARS M(pro)) dimer is the main functional form has been confirmed by experiment. However, because of the absence of structural information of the monomer, the reasons for this remain unknown. To investigate it, two molecular dynamics (MD) simulations in water for dimer and monomer models have been carried out, using the crystal structure of protomer A of the dimer as the starting structure for the monomer. During the MD simulation of dimer, three interest phenomena of protomer A have been observed: (i) the distance between NE2 of His41 and SG of Cys145 averages 3.72 A, which agrees well with the experimental observations made by X-ray crystallography; (ii) His163 and Glu166 form the "tooth" conformational properties, resulting in the specificity for glutamine at substrate P1 site; and (iii) the substrate-binding pocket formed by loop 140-146 and loop 184-197 is large enough to accommodate the substrate analog. However, during the MD simulation of the monomer complex, the three structural characteristics are all absent, which results directly in the inactivation of the monomer. Throughout the MD simulation of the dimer, the N-terminus of protomer B forms stable hydrogen bonds with Phe140 and Glu166, through which His163, Glu166, and loop 140-146 are kept active form. Furthermore, a water-bridge has been found between the N-terminus of protomer B and Gly170, which stabilizes His172 and avoids it moving toward Tyr161 to disrupt the H-bond between Tyr161 and His163, stabilizing the conformation of His163. The interactions between the N-terminus and another monomer maintain the activity of dimer.     

The periplasmic bacterial molecular chaperone SurA adapts its structure to bind peptides in different conformations to assert a sequence preference for aromatic residues

The periplasmic molecular chaperone protein SurA facilitates correct folding and maturation of outer membrane proteins in Gram-negative bacteria. It preferentially binds peptides that have a high fraction of aromatic amino acids. Phage display selections, isothermal titration calorimetry and crystallographic structure determination have been used to elucidate the basis of the binding specificity. The peptide recognition is imparted by the first peptidyl-prolyl isomerase (PPIase) domain of SurA. Crystal structures of complexes between peptides of sequence WEYIPNV and NFTLKFWDIFRK with the first PPIase domain of the Escherichia coli SurA protein at 1.3 A resolution, and of a complex between the dodecapeptide and a SurA fragment lacking the second PPIase domain at 3.4 A resolution, have been solved. SurA binds as a monomer to the heptapeptide in an extended conformation. It binds as a dimer to the dodecapeptide in an alpha-helical conformation, predicated on a substantial structural rearrangement of the SurA protein. In both cases, side-chains of aromatic residues of the peptides contribute a large fraction of the binding interactions. SurA therefore asserts a recognition preference for aromatic amino acids in a variety of sequence configurations by adopting alternative tertiary and quaternary structures to bind peptides in different conformations.     

How hormone receptor-DNA binding affects nucleosomal DNA: the role of symmetry.

Molecular dynamics simulations have been employed to determine the optimal conformation of an estrogen receptor DNA binding domain dimer bound to a consensus response element, ds(AGGTCACAGTGACCT), and to a nonconsensus response element, ds(AGAACACAGTGACCT). The structures simulated were derived from a crystallographic structure and solvated by a sphere (45-A radius) of explicit water and counterions. Long-range electrostatic interactions were accounted for during 100-ps simulations by means of a fast multipole expansion algorithm combined with a multiple time-step scheme in the molecular dynamics package NAMD. The simulations demonstrate that the dimer induces a bent and underwound (10.7 bp/turn) conformation in the DNA. The bending reflects the dyad symmetry of the receptor dimer and can be described as an S-shaped curve in the helical axis of DNA when projected onto a plane. A similar bent and underwound conformation is observed for nucleosomal DNA near the nucleosome's dyad axis that reflects the symmetry of the histone octamer. We propose that when a receptor dimer binds to a nucleosome, the most favorable dimer-DNA and histone-DNA interactions are achieved if the respective symmetry axes are aligned. Such positioning of a receptor dimer over the dyad of nucleosome B in the mouse mammary tumor virus promoter is in agreement with experiment.     

Crystal structures of monomeric actin bound to cytochalasin D

The fungal toxin cytochalasin D (CD) interferes with the normal dynamics of the actin cytoskeleton by binding to the barbed end of actin filaments. Despite its widespread use as a tool for studying actin-mediated processes, the exact location and nature of its binding to actin have not been previously determined. Here we describe two crystal structures of an expressed monomeric actin in complex with CD: one obtained by soaking preformed actin crystals with CD, and the other obtained by cocrystallization. The binding site for CD, in the hydrophobic cleft between actin subdomains 1 and 3, is the same in the two structures. Polar and hydrophobic contacts play equally important roles in CD binding, and six hydrogen bonds stabilize the actin-CD complex. Many unrelated actin-binding proteins and marine toxins target this cleft and the hydrophobic pocket at the front end of the cleft (viewing actin with subdomain 2 in the upper right corner). CD differs in that it binds to the back half of the cleft. The ability of CD to induce actin dimer formation and actin-catalyzed ATP hydrolysis may be related to its unique binding site and the necessity to fit its bulky macrocycle into this cleft. Contacts with residues lining this cleft appear to be crucial to capping and/or severing. The cocrystallized actin-CD structure also revealed changes in actin conformation. An ∼ 6° rotation of the smaller actin domain (subdomains 1 and 2) with respect to the larger domain (subdomains 3 and 4) results in small changes in crystal packing that allow the D-loop to adopt an extended loop structure instead of being disordered, as it is in most crystal structures of actin. We speculate that these changes represent a potential conformation that the actin monomer can adopt on the pathway to polymerization or in the filament.     

Solution structure of the disulfide-linked dimer of human intestinal trefoil factor (TFF3): the intermolecular orientation and interactions are markedly different from those of other dimeric trefoil proteins

The trefoil protein TFF3 forms a homodimer (via a disulfide linkage) that is thought to have increased biological activity over the monomer. The solution structure of the TFF3 dimer has been determined by NMR and compared with the structure of the TFF3 monomer and with other trefoil dimer structures (TFF1 and TFF2). The most significant structural differences between the trefoil domain in the monomer and dimer TFF3 are in the orientations of the N-terminal 3(10)-helix (residues 10-12) and in the presence in the dimer of an additional 3(10)-helix (residues 53-55) outside of the core region. The TFF3 dimer forms a more compact structure as compared with the TFF1 dimer where the two trefoil domains are connected by a flexible region with the monomer units being at variable distances from each other and in many different orientations. Although TFF2 is also a compact structure, the dispositions of its monomer units are very different from those of TFF3. The structural differences between the dimers result in the two putative receptor/ligand binding sites that remain solvent exposed in the dimeric structures having very different dispositions in the different dimers. Such differences have significant implications for the mechanism of action and functional specificity for the TFF class of proteins.     

Crystal structure of the human protein kinase CK2 regulatory subunit reveals its zinc finger-mediated dimerization.

Protein kinase CK2 is a tetramer composed of two alpha catalytic subunits and two beta regulatory subunits. The structure of a C-terminal truncated form of the human beta subunit has been determined by X-ray crystallography to 1.7 A resolution. One dimer is observed in the asymmetric unit of the crystal. The most striking feature of the structure is the presence of a zinc finger mediating the dimerization. The monomer structure consists of two domains, one entirely alpha-helical and one including the zinc finger. The dimer has a crescent shape holding a highly acidic region at both ends. We propose that this acidic region is involved in the interactions with the polyamines and/or catalytic subunits. Interestingly, conserved amino acid residues among beta subunit sequences are clustered along one linear ridge that wraps around the entire dimer. This feature suggests that protein partners may interact with the dimer through a stretch of residues in an extended conformation.     

Transmembrane helix-helix interactions: comparative simulations of the glycophorin a dimer

The glycophorin helix dimer is a paradigm for the exploration of helix-helix interactions in integral membrane proteins. Two NMR structures of the dimer are known, one in a detergent micelle and one in a lipid bilayer. Multiple (4 x 50 ns) molecular dynamics simulations starting from each of the two NMR structures, with each structure in either a dodecyl phosphocholine (DPC) micelle or a dimyristoyl phosphatidylcholine (DMPC) bilayer, have been used to explore the conformational dynamics of the helix dimer. Analysis of the helix-helix interaction, mediated by the GxxxG sequence motif, suggests convergence of the simulations to a common model. This is closer to the NMR structure determined in a bilayer than to micelle structure. The stable dimer interface in the final simulation model is characterized by (i) Gly/Gly packing and (ii) Thr/Thr interhelix H-bonds. These results demonstrate the ability of extended molecular dynamics simulations in a lipid bilayer environment to refine membrane protein structures or models derived from experimental data obtained in protein/detergent micelles.     

A molecular dynamics study of the C-terminal fragment of the L7/L12 ribosomal protein

The crystallographic dimer of the C-terminal fragment (CTF) of the L7/L12 ribosomal protein has been subjected to molecular dynamics (MD) simulations. A 90 picosecond (ps) trajectory for the protein dimer, 19 water molecules and two counter ions has been calculated at constant temperature. Effects of intermolecular interactions on the structure and dynamics have been studied. The exact crystallographic symmetry is lost and the atomic fluctuations differ from one monomer to the other. The average MD structure is more stable than the X-ray one, as judged by accessible surface area and energy calculations. Crystal (non-dimeric) interactions have been simulated in another 40 ps trajectory by using harmonic restraints to represent intermolecular hydrogen bonds. The conformational changes with respect ot the X-ray structure are then virtually suppressed.The unrestrained dimer trajectory has been scanned for cooperative motions involving secondary structure elements. The intrinsic collective motions of the monomer are transmitted via intermolecular contacts to the dimer structure.The existence of a stable dimeric form of CTF, resembling the crystallographic one, has been documented. At the cost of fairly small energy expenditure the dimer has considerable conformational flexibility. This flexibility may endow the dimer with some functional potential as an energy transducer.