David R. Langley*
Bristol-Myers Squibb Company,
Pharmaceutical Research Institute,
5 Research Parkway, P.O. Box 5100,
Wallingford, CT 06492-7660, USA
*For author correspondence. Phone: 203-284-6656; Fax: 203-284-7702; E-mail:
david@viper.wfd.pri.bms.com
Abstract
Molecular dynamic (MD) simulations using the BMS nucleic acid force field produce environment and sequence dependent DNA conformations that closely mimic experimentally derived structures. The parameters were initially developed to reproduce the potential energy surface, as defined by quantum mechanics, for a set of small molecules that can be used as the building blocks for nucleic acid macromolecules (dimethyl phosphate, cyclopentane, tetrahydrofuran, etc.). Then the dihedral parameters were fine tuned using a series of condensed phase MD simulations of DNA and RNA (in zero added salt, 4M NaCl, and 75% ethanol solutions). In the tuning process the free energy surface for each dihedral was derived from the MD ensemble and fitted to the conformational distributions and populations observed in 87 A- and B-DNA x-ray and 17 B-DNA NMR structures. Over 41 nanoseconds of MD simulations are presented which demonstrate that the force field is capable of producing stable trajectories, in the correct environments, of A-DNA, double stranded A-form RNA, B-DNA, Z-DNA, and a netropsin-DNA complex that closely reproduce the experimentally determined and/or canonical DNA conformations. Frequently the MD averaged structure is closer to the experimentally determined structure than to the canonical DNA conformation. MD simulations of A- to B- and B- to A-DNA transitions are also shown. A-DNA simulations in a low salt environment cleanly convert into the B-DNA conformation and converge into the RMS space sampled by a low salt simulation of the same sequence starting from B-DNA. In MD simulations using the BMS force field the B-form of d(GGGCCC)2 in a 75% ethanol solution converts into the A-form. Using the same methodology, parameters, and conditions the A-form of d(AAATTT)2 correctly converts into the B-DNA conformation. These studies demonstrate that the force field is capable of reproducing both environment and sequence dependent DNA structures. The 41 nanoseconds (nsec) of MD simulations presented in this paper paint a global picture which suggests that the DNA structures observed in low salt solutions are largely due to the favorable internal energy brought about by the nearly uniform screening of the DNA electrostatics. While the conformations sampled in high salt or mixed solvent environments occur from selective and asymmetric screening of the phosphate groups and DNA grooves, respectively, brought about by sequence induced ion and solvent packing.
M. Ravi KiranÝ and Manju Bansal*
Molecular Biophysics Unit, Indian Institute of Science, Bangalore -
560 012, India
ÝPresent Address: National Centre for Biological
Sciences, P.O. Box 1234, I.I.Sc. Campus, Bangalore 560 012, India
*Author to whom correspondence should be addresed. Phone: 91-80-3092534;
Fax: 91-80-3341683/3348535; E-Mail: mb@mbu.iisc.ernet.in
Abstract
Molecular dynamics (MD) studies have been carried out on the Hoogsteen hydrogen bonded parallel and the reverse Hoogsteen hydrogen bonded antiparallel C.G*G triplexes. Earlier, the molecular mechanics studies had shown that the parallel structure was energetically more favourable than the antiparallel structure. To characterize the structural stability of the two triplexes and to investigate whether the antiparallel structure can transit to an energetically more favourable structure, due to the local fluctuations in the structure during the MD simulation, the two structures were subjected to 200ps of constant temperature vacuum MD simulations at 300K. Initially no constraints were applied to the structures and it was observed that for the antiparallel triplex, the structure showed a large root mean square deviation from the starting structure within the first 12ps and the N4-H41--O6 hydrogen bond in the WC duplex got distorted due to a high propeller twist and a moderate increase in the opening angle in the basepairs. Starting from an initial value of 30°, helical twist of the average structure from this simulation had a value of 36°, while the parallel structure stabilized at a twist of 33°. In spite of the hydrogen bond distortions in the antiparallel triplex, it was energetically comparable to the parallel triplex. To examine the structural characteristics of an undistorted structure, another MD simulation was performed on the antiparallel triplex by constraining all the hydrogen bonds. This structure stabilized at an average twist of 33°. In the course of the dynamics though the energy of the molecule - compared to the initial structure - improved, it did not become comparable to the parallel structure. Energy minimization studies performed in the presence of explicit water and counterions also showed the two structures to be equally favourable energetically. Together these results indicate that the parallel C.G*G triplex with Hoogsteen hydrogen bonds also represents a stereochemically and energetically favourable structure for this class of triplexes.
Sukesh R. Bhaumik, Kandala V.R. Chary* and Girjesh Govil
Department of Chemical Sciences, Tata Institute of Fundamental Research,
Homi Bhabha Road, Mumbai 400 005, India
*Author to whom correspondence may be addressed. Phone: 215 2971/2979; Fax:
091-22-215 2110/2181; E-mail: Chary@tifr.res.in
Abstract
We have carried out molecular modeling of a triple stranded pyrimidine(Y) . purine(R) : pyrimidine(Y) (where ':' refers to Watson-Crick and '.' to Hoogsteen bonding) DNA, formed by a homo-purine (d-TGAGGAAAGAAGGT) and homo-pyrimidine (d-CTCCTTTCTTCC). Molecular mechanics calculations using NMR constraints have provided a detailed three dimensional structure of the triplex. The entire stretches of purine and the pyrimidine nucleotides have a conformation close to B-DNA. The three strands are held by the canonical C+.G:C and T.A:T hydrogen bonds. The structure also contains two mismatch C+.G-T and T.A+-C base triples which have been characterized for the first time. In the A+-C base-pair of the T.A+-C triple, both hydrogen donors are situated on the purine (A+(1N) and A+(6N)). We observe a unique hydrogen bonding interaction scheme in case of C+.G-T where one acceptor, G(6O), is bonded to three donors (C+(3NH), C+(4NH2) and T(3NH)). Though the C+.G-T base triple is less stable than C+.G:C, it is significantly more stable than T.A:T. On the other hand, T.A+-C is as stable as the T.A:T base triad.
Guillaume Bertucat, Richard Lavery and Chantal Prévost*
Laboratoire de Biochimie Théorique, UPR 9080, Institut de Biologie
Physico-Chimique, 13, rue Pierre et Marie Curie, 75005 Paris, France
*Author to whom correspondence should be addressed. Phone: 33 1 43 25 26
09; Fax: 33 1 40 46 83 31; E-mail: Chantal.Prevost@ibpc.fr
Abstract
The nucleoproteic filaments of RecA polymerized on single stranded DNA are able to integrate double stranded DNA in a coaxial arrangement (with DNA stretched by a factor 1.5), to recognize homologous sequences in the duplex and to perform strand exchange between the single stranded and double stranded molecules. While experimental results favor the hypothesis of an invasion of the minor groove of the duplex by the single strand, parallel minor groove triple helices have never been isolated or even modeled, the minor groove offering little space for a third strand to interact. Based on an internal coordinate modeling study, we show here that such a structure is perfectly conceivable when the two interacting oligomers are stretched by a factor 1.5, in order to open the minor groove of the duplex. The model helix presents characteristics that coincide with known experimental data on unwinding, base pair inclination and inter-proton distances. Moreover, we show that extension and unwinding stabilize the triple helix. New patterns of triplet interaction via the minor groove are presented.
Aleksej Yu. Denisov1, E.V. Zamaratski1, Tatiana V. Maltseva1,
Anders Sandström1, Somer Bekiroglu1, Karl-Heinz Altmann2, Martin
Egli3 and Jyoti Chattopadhyaya1*
1Department of Bioorganic Chemistry, Box 581, Biomedical
Center, University of Uppsala, S-751 23 Uppsala, Sweden
2Department of Oncology, Novartis Pharma Inc.,
CH-4002 Basel, Switzerland
3Drug Discovery Program, Department of Molecular
Pharmacology and Biological Chemistry, Northwestern University Medical School,
Chicago, IL 60611-3008, USA
Summary
The NMR conformation of a carbocyclic analog of the Dickerson-Drew dodecamer [d(CGCGAAT*T*CGCG)]2 containing 6'-alpha-Me carbocyclic thymidines (T*) has been determined and compared with that of its X-ray structure. The solution structure of the 6'-alpha-Me carbocyclic thymidine modified duplex has also been compared with the solution structure of the corresponding unmodified Dickerson-Drew duplex solved by us under the same experimental conditions. The NMR structures have been based on 24 experimental distance and torsion constraints per residue for [d(CGCGAAT*T*CGCG)]2 (1) and on 21 constraints per residue for the natural counterpart. In general, both final NMR structures are more close to the B-type DNA. The cyclopentane moieties of the carbocyclic thymidine residues adopt C1'-exo B-DNA type puckers (the phase angles P = 136-139° and the puckering amplitudes Y = 36-37°) that are close to their previously published crystal C1'-exo or C2'-endo puckers. The main differences between the two NMR structures are for beta(T*8) and epsilon, x(T*7) backbone torsions (27-50°), for basepair twist for the 7-8 and 8-9 basepair steps (5-6°), tilt for the 8-9 step (7°), roll for the 7-8 step (7°), shift for the 7-8 step (0.9Å) and slide for the 9-10 step (0.6Å). The relatively small deviations of helical structure parameters lead to structural isomorphism of these duplexes in aqueous solutions (atomic RMSD = 1.0Å). The difference of the minor groove widths (less than 0.7Å) in the core part of the modified duplex in comparison with the native one is much smaller than the difference between the X-ray structures of these duplexes. A detailed comparison of NMR and X-ray structure parameters showed significant monotonic differences (0.9-2.5Å) for all basepair slides in both duplexes. Deviations between NMR and X-ray structure parameters for the modified duplex were also found for basepair tilt of the 4-5 step (13°), rolls for the 8-9 and 10-11 steps (16° and 19°), twist of the 3-4 step (8°) and shift of the 9-10 step (0.9Å).
T. V. Maltseva1, K.-H. Altmann2,
M. Egli3 and J. Chattopadhyaya1*
1Department of Bioorganic Chemistry, Box 581, Biomedical
Center, University of Uppsala,S-751 23 Uppsala, Sweden
2Department of Oncology, Novartis Pharma Inc,
Bldg. K136.421,CH-4002 Basel, Switzerland
3Drug Discovery Program and Department of Molecular
Pharmacology and Biological Chemistry, Northwestern University Medical School,
Chicago, IL 60611-3008, USA
*Author to whom correspondence should be addressed. Phone: +46-18-4714577;
Fax: +46-18-554495; E-mail: jyoti@bioorgchem.uu.se.
Summary
The residence time of the bound water molecules in the antisense oligodeoxyribonucleotides containing 7'-alpha-methyl (TMe) carbocyclic thymidines in duplex (I), d5'(1C2G3C4G5A6A7TMe8TMe9C10G11C12G)23', and 6'-alpha-hydroxy (TOH) carbocyclic thymidines in duplex (II), d5'(1C2G3C4G5AOH6AOH7TOH8 TOH9C10G11C12G)23, have been investigated using a combination of NOESY and ROESY experiments. Because of the presence of 7'-alpha-methyl groups of TMe in the centre of the minor groove in duplex (I), the residence time of the bound water molecule is shorter than 0.3 ns. The dramatic reduction of the residence time of the water molecule in the minor groove in duplex (I) compared with the natural counterpart has been attributed to the replacement of second shell of hydration and disruption of hydrogen-bonding with O4' in the minor groove by hydrophobic alpha-methyl groups, as originally observed in the X-ray study. This effect could not be attributed to the change of the width of the minor groove because a comparative NMR study of the duplex (I) and its natural counterpart showed that the widths of their minor grooves are more or less unchanged (r.m.s.d change in the core part is <0.63Å). For duplex (II) with polar 6'-alpha-hydroxyl groups pointed to the minor groove, the correlation time is much longer than 0.36ns as a result of the stabilising hydrogen-bonding interaction with N3 or O2 of the neighbouring nucleotides.
A.P. LyubartsevÝ and A. Laaksonen*
Division of Physical Chemistry, Arrhenius Laboratory, Stockholm University,S-106
91, Stockholm, Sweden
ÝAlso affiliated with: Institute of Physics,
St. Petersburg State University, 198904 St. Petersburg, Russia
*Author to whom correspondence should be addressed. Phone: +46-8-162372,
Fax: +46-8-152187, E-mail: aatto@physc.su.se
Abstract
Molecular dynamics simulations of the [d(ATGCAGTCAG]2 fragment of DNA, in water and in the presence of three different counter-ions (Li+, Na+ and Cs+) are reported. Three-dimensional hydration structure and ion distribution have been calculated using spatial distribution functions for a detailed picture of local concentrations of ions and water molecules around DNA. According to the simulations, Cs+ ions bind directly to the bases in the minor groove, Na+ ions bind prevailing to the bases in the minor groove through one water molecule, whereas Li+ ions bind directly to the phosphate oxygens. The different behavior of the counter-ions is explained by specific hydration structures around the DNA and the ions. It is proposed how the observed differences in the ion binding to DNA may explain different conformational behavior of DNA. Calculated self-diffusion coefficients for the ions agree well with the available NMR data.
Anne Lebrun¦ and Richard Lavery*
Laboratoire de Biochimie Théorique, Institut de Biologie Physico-Chimique,
13, rue Pierre et Marie Curie, 75005 Paris, France
¦Present address: Department of Physiology
and Biophysics, Mount Sinai School of Medicine, New York, New York 10029,
USA
*Author to whom correspondence should be addressed. Phone: 33 1 43 25 26
09; Fax: 33 1 43 29 56 45; E-mail: richard@ibpc.fr
Abstract
DNA stretching and strand separation have been studied by molecular mechanics using an oligomer which has been the subject of nanomanipulation experiments (Noy et al., Chem. Biol. 4, 519, 1997). Adiabatic mapping of conformational energy carried out as a function of stretching leads to force/extension curves in good correlation with the experimental results. Other types of deformation are also modeled and compared with the experimental results obtained on polymeric DNA. The results highlight overall similarities, but point to thermodynamic differences and also to local base sequence effects which can be expected to play an important role at the level of biologically induced structural deformations.
Raphael Gurlie and Krystyna Zakrzewska*
Laboratoire de Biochimie Théorique, UPR 9080 CNRS, Institut de
Biologie Physico-Chimique 13, rue Pierre et Marie Curie, 75005 Paris, France
*Author to whom correspondence should be addressed. Phone: 33 1 43 25 26
09; Fax: 33 1 43 29 56 45
Abstract
A theoretical study is presented of the influence of salt bridges between protein cationic side chains and DNA phosphates on DNA conformation and flexibility. Two DNA sequences are studied containing respectively the HNF3 and CAP binding sites. The effect of salt bridges is modelled by the neutralisation of net phosphate charges for the groups involved in such interactions in the complex. Energy optimised conformations are obtained by molecular mechanics calculations using the JUMNA program. Base sequence dependence is studied by moving the phosphate neutralisation pattern along the sequence, while normal mode analysis is used to evaluate DNA flexibility. The results show that phosphate neutralisation has a strong influence on DNA conformation. For the HNF3 binding sequence, the free oligomer is bent in direction very different from that observed in the protein complex. Phosphate neutralisation changes this direction by 45° to within only 4° of the direction in the complex, without changing the bending angle. For the CAP binding sequence, the free oligomer is already intrinsically curved in the direction favoured by the protein, but phosphate neutralisation increases the bending angle. For both oligomers studied these effects are strongly sequence dependent. It is also shown that oligomer flexibility cannot be explained by a simple superposition of the properties of successive dinucleotide steps. Important long range coupling effects are observed. However, for both sequence studied, phosphate neutralisation however leads to a reduction in oligomer flexibility.
Julio C. Facelli*
Center for High Performance Computing, University of Utah, Salt Lake
City, UT 84112-0190
*For author correspondence. Phone (801)-581-7529; Fax: (801)-585-5366; E-mail:
facelli@chpc.utah.edu
Abstract
This paper presents ab initio (DFT) calculations of the 15N chemical shifts in AT (Adenine-Thymine) and CG (Cytosine-Guanine) nucleic acid base pairs. Calculations were done on 14 AT and 18 CG base pairs using experimental (X-ray) geometries obtained from several DNA decamers. The calculated chemical shifts are compared with the experimental values in the pure bases and subjected to statistical analysis to explore their sensitivity to the local geometry and pair helix parameters. The results indicate that the 15N chemical shifts, isotropic and principal components are quite sensitive to small changes in the geometry of the pairs, but they do not correlate well with the helix pair parameters. From the statistical analysis, several linear correlations between structural parameters and chemical shifts emerge. These relationships may serve as a foundation to extract information on molecular structure from 15N chemical shift measurements.
Michael Montrel1*, Vasily P. Chuprina2,3, Valery I. Poltev1,4,
Willy Nerdel5 and Einar Sletten5
1Institute of Theoretical and Experimental Biophysics, Russian
Academy of Sciences, 142292 Pushchino, Moscow Region, Russia
2Institute of Mathematical Problems of Biology,
Russian Academy of Sciences, Pushchino, Moscow Region, Russia
3Bijvoet Center of Biomolecular Research, Utrecht,
Netherlands
4Facultad de ciencias fisico matematicas, Benemerito
universidad autonomo de Puebla, Mexico
5Department of Chemistry, University of Bergen, N-5007 Bergen, Norway
*Author to whom correspondence should be addressed. Phone: +7(0967) 739-205;
Fax: +7(0967) 790-553; E-mail: montrel@venus.iteb.serpukhov.su
Abstract
Experimentally observed sequence-selective binding of metal ion to DNA oligonucleotides have been compared with variations of electrostatic potential (EP) along the helix. Calculations of EP have been performed for three atomic models of the oligonucleotide duplex [d(CGCGAATTCGCG)2] using several variants of EP calculations, including a solution of non-linear Poisson-Boltzmann equation (NPBE). N7 atom of guanine adjacent to adenine base was identified as a region with the most negative electrostatic potential in the major groove. The EP value for the Me ion binding site surpasses the value for N7 of other guanines by 10-26% depending on particular duplex conformation. Qualitatively, the sequence dependent variations of EP near guanine N7 atoms are in agreement with the sequence-selective behavior of Mn(II) and Zn(II) ions as revealed by NMR experiments. But the difference in EP between the two most negative regions near guanine N7 atoms does not exceed 1.25 kT/e. Simple model suggests that metal ions are capable to form ion-hydrate complexes with G-Pu steps of DNA duplex. These complexes are formed via one Me...G and five Me...water coordination bonds with water molecules hydrogen bonded to two adjacent purine bases in the same chain. We suppose that such a stereospecific structural possibility is the main factor which control the sequence-selectivity in the metal ion binding. A combination of both mechanisms allows to explain sequence specific Mn(II) and Zn(II) binding to a set of oligonucleotides.
S. Lepore1, O. Mauffret1,
S. El Antri2, O. Convert3,
E. Lescot1, G. Tevanian1
and D. Fermandjian1*
1Département de Biologie Structurale UMR
1772 CNRS, Institut Gustave Roussy39-53, rue Camille Desmoulins94805 Villejuif,
France
2Laboratoire de Chimie Bioorganique et Analytique
Faculté des Sciences et Techniques,BP 146 Mohammedia, MAROC
3Laboratoire de Chimie Structurale Organique
et Biologique Université Pierre et Marie CurieUMR 7613 CNRS, Bat.
F 8, rue Cuvier 75005 Paris, France
*Author to whom correspondence should be addressed. Phone: 33.(1).42.11.49.85;
Fax: 33.(1).42.11.52.76; E-mail: sfermand@igr.fr
Abstract
The hydration properties of the non-palindromic duplex d(CTACTGCTTTAG). d(CTAAAGCAGTAG) were investigated by NMR spectroscopy. The oligonucleotide possesses a heterogeneous B-DNA structure. The H2(n)-H1'(m+1) distances reflect a minor groove narrowing within the TTT/AAA segment (~ 3.9Å) and a sudden widening at the T10:A15 base-pair (~ 5.3Å) , the standard B-DNA distance being ~ 5Å. The facing T10pA11 and T14pA15 steps at the end of the TTTA/AAAT segment have completely different behaviors. Only A15 ending the AAA run displays NMR features comparable to those shown by adenines of TpA steps occupying the central position of TnAn (n > 2) segments. These involve particular chemical shifts and line broadening of the H2 and H8 protons. Positive NOESY cross-peaks were measured between the water protons and the H2 protons of A15, A16 and A17 reflecting the occurrence of hydration water molecules with residence times longer than 500 picoseconds along the minor groove of the TTT/AAA segment. In contrast no water molecules with long residence times were observed neither for A3, A20 and A23 nor for A11 ending the 5'TTTA run. We confirm thus that the binding of water molecules with long residence time to adenine residues correlates with the minor groove narrowing. In contrast, the widening of the minor groove at the A11:T14 base-pair ending the TTTA/TAAA segment, likely associated to a high negative propeller twist value at this base-pair, prevents the binding of a water molecule with long residence time to A11 but not to A15 of the preceding T10:A15 base-pair. Thus, in our non-palindromic oligonucleotide the water molecules bind differently to A11 and A15 although both adenines are part of a TpA step. The slower motions occurring at A15 compared to A11 are also well explained by the present results.
Alex Ninabar and Julia M. Goodfellow*
Department of Crystallography, Birkbeck College, University of London,
Malet Street, London WC1E 7HX, UK
*Author to whom correspondence should be addressed. Phone: 44 171 631 6800;
Fax: 44 171 631 6803; E-mail: j.goodfellow@mail.cryst.bbk.ac.uk
Abstract
DNA damage produced by free radicals is probably the most frequent lesion encountered by cells (Wallace, S.S., Environmental and Molecular Mutagenesis 12, 431-477, 1988 (1)). One of the most common effects is the formation of 7-hydro-8-oxodeoxyguanine due to oxygen radicals interacting with the normal guanine base. Such chemical changes appear to be important in mutagenesis, cancer and aging. We have used computer simulation techniques to model the effect of inclusion of such a modified base within a duplex strand of DNA. We find that such modifications can be stabilized within a normal sequence. The conformation of the modified base relative to the sugar residue depends on many local interactions not accessible to the isolated nucleoside. We have also studied the essential dynamics of both normal and modified sequences and show that there are only subtle changes to the dynamics on inclusion of such a modification.
Hong Qian1* and James White2
1Department of Applied Mathematics, University of Washington,
Seattle, WA 98195
2Department of Mathematics, University of California,
Los Angeles, CA 90095
*Author to whom correspondence should be addressed. Phone: 206-685-2971;
Fax: 206-685-1440; Email: qian@amath.washington.edu
Abstract
It is well known that a large linking number induces an abrupt writhing of a circular rod with zero intrinsic curvature, i.e., the stress-free state of the rod is straight. We show here that for any rod with a uniform natural curvature, no matter how small the intrinsic curvature is, a twist will induce a continuous writhing from the circular configuration and the abrupt writhing is only the limiting case when the intrinsic curvature is absolutely zero. The implication of this result on elastic models of circular DNA is discussed.
Jayashree Srinivasan1, Jennifer Miller2, Peter A. Kollman2 and
David A. Case1*
1Department of Molecular Biology, The Scripps Research
Institute, La Jolla, CA 92037
2Department of Pharmaceutical Chemistry, University
of California, San Francisco, CA 94143
*Author to whom correspondence should be addressed. Phone: 619-784-9768:
Fax: 619-784-8896; E-mail: case@scripps.edu.
Abstract
We apply continuum solvent models to investigate the relative stability of various conformational forms for two RNA sequences, GGAC(UUCG)GUCC and GGUG(UGAA)CACC. In the first part, we compare alternate hairpin conformations to explore the reliability of these models to discriminate between different local conformations. A second part looks at the hairpin-duplex conversion for the UUCG sequence, identifying major contributors to the thermodynamics of a much large scale transition. Structures were taken as snapshots from multi-nanosecond molecular dynamics simulations computed in a consistent fashion using explicit solvent and with long-range electrostatics accounted for using the Particle-Mesh Ewald procedure. The electrostatic contribution to solvation energies were computed using both a finite-difference Poisson-Boltzmann (PB) model and a pairwise Generalized Born model; non-electrostatic contributions were estimated with a surface-area dependent term. To these solvation free energies were added the mean solute internal energies (determined from a molecular mechanics potential) and estimates of the solute entropy (from a harmonic analysis). Consistent with experiment and with earlier solvated molecular dynamics simulations, the UUCG hairpin was found to prefer conformers close to a recent NMR structure determination in preference to those from an earlier NMR study. Similarly, results for the UGAA hairpin favored an NMR-derived structure over that to be expected for a generic GNRA hairpin loop. Experimental free energies are not known for the hairpin/duplex conversion, but must be close to zero since hairpins are seen in solution and duplexes in crystals; out calculations find a value near zero and illustrate the expected interplay of solvation, salt effects and entropy in affecting this equilibrium.
Michael J. Seewald, Armin U. MetzgerÝ,
Dieter Willbond, Paul Rösch and Heinrich Sticht*
Lehrstuhl für Biopolymere der Universität Bayreuth, Universitätsstr.
30, 95447 Bayreuth, Germany
ÝPresent address: Merck KGaA, Frankfurter
Str. 250, 64293 Darmstadt, Germany
*Author to whom correspondence should be addressed. Phone: +49 921 553542;
Fax: +49 921 553544; E-mail: Heinrich.Sticht@Uni-Bayreuth.de
Abstract
The trans-activator protein (Tat) of human immunodeficiency virus type 1 (HIV-1) binds to an uridine-rich bulge of an RNA target (TAR; trans-activation responsive element) predominantly via its basic sequence domain. The structure of the Tat(46-58)-TAR complex has been determined by a novel modeling approach relying on structural information about one crucial arginine residue and crosslink data. The strategy described here solely uses this experimental data without additional "modeling" assumptions about the structure of the complex in order to avoid human bias. Model building was performed in a fashion similar to structure calculations from nuclear magnetic resonance (NMR)-spectroscopic data using restrained molecular dynamics.
The resulting set of structures of Tat(46-58) in its complex with TAR reveals that all models have converged to a common fold, showing a backbone root mean square deviation (RMSD) of 1.36Å. Analysis of the calculated structures suggests that HIV-1 Tat forms a hairpin loop in its complex with TAR that shares striking similarity to the hairpin formed by the structure of the bovine immunodeficiency virus Tat protein after TAR binding as determined by NMR studies. The outlined approach is not limited to the Tat-TAR complex modeling, but is also applicable to all molecular complexes with sufficient biochemical and biophysical data available.
Pascal Auffinger and Eric Westhof*
Institut de Biologie Moléculaire et Cellulaire du CNRS, Modélisations
et Simulations des Acides Nucléiques, UPR 9002, 15 rue René
Descartes, 67084 Strasbourg Cedex, France
*Author to whom correspondence should be addressed. Phone: +33 (0)3 88.41.70.00;
Fax: +33 (0)3 88.41.70.46; E-mail: westhof@ibmc.u-strasbg.fr
Abstract
The hydration patterns around the RNA Watson-Crick and non-Watson-Crick base pairs in crystals are analyzed and described. The results indicate that (i) the base pair hydration is mostly "in-plane"; (ii) eight hydration sites surround the Watson-Crick G-C and A-U base pairs, with five in the deep and three in the shallow groove, an observation which extends the characteristic isostericity of Watson-Crick pairs; (iii) while the hydration around G-C base pairs is well defined, the hydration around A-U base pairs is more diffuse; (iv) the hydration sites close to the phosphate groups are the best defined and the most recurrent ones; (v) a string of water molecules links the two shallow groove 2'-hydroxyl groups, and (vi) the water molecules fit into notches, the size and accessibility of which are almost as important as the number and strength of the hydrophilic groups lining the cavity. Residence times of water molecules at specific hydration sites, inferred from molecular dynamics simulations, are discussed in the light of present data.
Margaret S. VanLoock, Brett A. Harris and Stephen C. Harvey*
Department of Biochemistry and Molecular Genetics, University of Alabama
at Birmingham, Birmingham, AL 35294-0005
*Author to whom correspondence should be addressed. Phone: (205) 934-5028;
Fax: (205) 975-2547; E-mail: harvey@uab.edu
Abstract
The presence of topological knots in large RNA structures is highly unlikely given that 1) no RNA structures determined thus far contain topological knots, 2) secondary structure maps for most RNA molecules are knot free, 3) there are no known RNA topoisomerases, and 4) it is difficult to imagine how knots could be formed specifically and uniquely during transcription. Since native RNA structures probably lack topological knots, models of these RNA molecules should be free of knots as well. Therefore, we have examined four existing models for the 30S ribosomal subunit to determine if any of the three domains of the 16S rRNA molecule is knotted. We found that all but one model had at least one knotted domain. We conclude that models of large RNA molecules should be examined for knotting before publication.
M.S. Madhusudhan and Saraswathi Vishveshwara*
Molecular Biophysics Unit, Indian Institute of Science, Bangalore -560
012, India
*Author to whom correspondence should be addressed. Phone: 91-80-3092611;
Fax: 91-80-3348535; E-mail: sv@mbu.iisc.ernet.in
Abstract
Angiogenin belongs to the Ribonuclease superfamily and has a weak enzymatic activity that is crucial for its biological function of stimulating blood vessel growth. Structural studies on ligand bound Angiogenin will go a long way in understanding the mechanism of the protein as well as help in designing drugs against it. In this study we present the first available structure of nucleotide ligand bound Angiogenin obtained by computer modeling. The importance of this study in itself notwithstanding, is a precursor to modeling a full dinucleotide substrate onto Angiogenin. Bovine Angiogenin, the structure of which has been solved at a high resolution, was earlier subjected to Molecular Dynamics simulations for a nanosecond. The MD structures offer better starting points for docking as they offer lesser obstruction than the crystal structure to ligand binding. The MD structure with the least serious short contacts was modeled to obtain a steric free Angiogenin - 3' mononucleotide complex structure. The structures were energetically minimized and subjected to a brief spell of Molecular Dynamics. The results of the simulation show that all the ligand-Angiogenin interactions and hydrogen bonds are retained, redeeming the structure and docking procedure. Further, following ligand - protein interactions in the case of the ligands 3'-CMP and 3'-UMP we were able to speculate on how Angiogenin, a predominantly prymidine specific ribonuclease prefers Cytosine to Uracil in the first base position.
Mihaly Mezei1* and Frank Guarnieri1,2
1Department of Physiology and Biophysics, Mount Sinai School
of Medicine, CUNY, New York, NY 10029
2Sarnoff Corporation, 201 Washington Road, Princeton,
NJ 08543
*Author to whom correspondence should be addressed. Phone: 212-241-2186;
Fax: 212-860-3369; E-mail: mezei@inka.mssm.edu
Abstract
The conformational preference of the gonadotropin-releasing hormone (GnRH) and its Lys-8 mutant, studied earlier with a continuum model, was revisited using an explicit solvent model and thermodynamic integration to calculate the solvents contribution to the conformation-dependence of its free energy. In addition, the Proximity Criterion was used to further analyze the effects of conformational changes.
Wolfgang Weber1, Hagop Demirdjian2,
Roberto D. Lins3, James M. Briggs1*,
Ricardo Ferreira3 and J. Andrew McCammon1
1Department of Pharmacology and Department of Chemistry and
Biochemistry, University of California, San Diego, La Jolla, CA 92093-0365,
USA
2On leave from: Departement des Sciences de
la Matiere, Ecole Normale Supérieure de Lyon, 46 allée d'Italie,
F-69364 Lyon, France
3On leave from: Departamento de Quimica Fundamental,
Universidade Federal de Pernambuco, Recife, PE 50670-901, Brazil
*Author to whom correspondence should be addressed. Phone: (713) 743-8366;
Fax: (713) 743-8351; E-mail: jbriggs@uh.edu.
Abstract
The three-dimensional structure of the active site region of the enzyme HIV-1 integrase is not unambiguously known. This region includes a flexible peptide loop that cannot be well resolved in crystallographic determinations. Here we present two different computional approaches with different levels of resolution and on different time-scales to understand this flexibility and to analyze the dynamics of this part of the protein. We have used molecular dynamics simulations with an atomic model to simulate the region in a realistic and reliable way for 1 ns. It is found that parts of the loop wind up after 300 ps to extend an existing helix. This indicates that the helix is longer than in the earlier crystal structures that were used as basis for this study. Very recent crystal data confirms this finding, underlining the predictive value of accurate MD simulations. Essential dynamics analysis of the MD trajectory yields an anharmonic motion of this loop. We have supplemented the MD data with a much lower resolution Brownian dynamics simulation of 600 ns length. It provides ideas about the slow-motion dynamics of the loop. It is found that the loop explores a conformational space much larger than in the MD trajectory, leading to a "gating"-like motion with respect to the active site.
Evans Coutinho1, Shantaram Kamath1,
Anil Saran2* and Sudha Srivastava2
1Bombay College of Pharmacy, Kalina, Mumbai 400 098, India
2Tata Institute of Fundamental Research, Homi
Bhabha Road, Navy Nagar, Mumbai 400 005, India
*Author to whom correspondence should be addressed. Phone: 2152971, Ext.
2377; Fax: 091-22-215 2110/2181; E-mail: anil@tifrvax.tifr.res.in.
Abstract
The conformation of the C-terminal octapeptide fragment of Substance P (SP4-11, Pro-Gln-Gln-Phe-Phe-Gly-Leu-Met-NH2) has been investigated by 2D-NMR and MD methods. The octapeptide exists in a blend of conformations. The molecule seems to shuttle between conformations with g-bends either at Phe5 or Gly6 or Gln3 or Leu7 and between a nearly extended structure.