Shu-Yun Le1, Jih-H. Chen2,
N. Pattabiraman2 and Jacob V. Maizel, Jr.1
1Laboratory of Experimental and Computational Biology,
DBS, FCRDC, National Cancer Institute, NIH, Frederick, MD 21701
2Frederick Biomedical Supercomputing Center,
SAIC-NCI/FCRDC, P.O. Box B, Fresderick, MD 21702
*Author to whom correspondence should be addressed. Phone: 301-846-5576;
Fax: 301-846-5598; E-mail: shuyun@orleans.ncifcrf.gov
Abstract
The three-dimensional (3-D) structure of a RNA pseudoknot that causes
the efficient ribosomal frameshifting in the gag-pro region of mouse mammary
tumor virus (MMTV) has been determined recently by nuclear magnetic resonance
(NMR) studies (9). But since the structure refinement in the studies did
not use metal ions and waters, it is not clear how metal ions participate
in the stabilization of the pseudoknot, and what kind of ion-RNA interactions
dominate in the tertiary contacts for the RNA pseudoknotting. Based on the
reported structure data of the pseudoknot VPK of MMTV (9), we gradually
refined the structure by restrained molecular dynamics (MD) using NMR distance
restraints (17). Restrained MD simulation of the RNA pseudoknot was performed
with sodium ions and water molecules. Our results are in good agreement
with known NMR data and delineate the importance of the metal ion coordination
in the stability of the pseudoknot. In the non-coaxially stacking pseudoknot,
stem 1 (S1), stem 2 (S2), and the intervening A14 involves unconventional
stacking of base pairs coordinated by Na+ and/or bridging water molecules.
A6 and G7 of loop L1 make a perfect base stacking in the major groove and
are further stabilized by coordinated Na+ ions
and water molecules. The first 4-nucleotide (nt) ACUC of loop L2 form a
sharp turn and the following 4-nt AAAA cross the minor groove of S1 and
are steadied by interactions with the nucleotides of S1, bridging water
molecules and coordinated Na+ ions. Our studies suggest that the
metal ion plays a crucial role in the RNA pseudoknotting of VPK. In the
stacking interior of S1 and S2, the Na+ ion is positioned in the
major groove and interacts directly with the carbonyl group O6 of G28 and carbonyl group O4
of U13 in the wobble base pair U13:G28. The ion-RNA interactions in MMTV
VPK not only stabilize the RNA pseudoknot but also modify the electrostatic
properties of the nucleotides at the critical parts of the pseudoknot VPK.
Carol A. Edwards1, Chang-Shung Tung2, Louis A. Silks3, Joe M.
Gatewood1, James A. Fee4
and S. V. Santhana Mariappan1*
1Structural Biology,
LS-8, MS M880,
2Theoretical Biology and Biophysics, T-10, MS
K710,
3Bioscience and Biotechnology, CST-4, MS C345,
Los Alamos National Laboratory, Los Alamos, NM 87545
and 4Dept. Biology, UCSD,
La Jolla, CA 92037
Abstract
In this paper, a general method is developed to study site-specific interactions in DNA-protein complexes using heteronuclear NMR spectroscopy and molecular modeling. This method involves two steps: (a) homonuclear 1H NMR and molecular modeling are used to develop a low resolution model and (b) 15N7-guanosine containing oligonucleotides are employed to probe the specific intermolecular interactions predicted in (a). This method is applied to Cro-operator complex due to its small size and extensive prior characterization. Non-exchangeable and exchangeable base protons have been assigned by nuclear Overhauser effect spectroscopy (NOESY) and chemical shift correlation spectroscopy. Extensive line-broadening has been observed in the 1H NMR spectra of the operator DNA in the presence of protein. Differential line-broadening observed in the imino proton region and the comparison of NOESY spectra in the presence and absence of Cro protein show that guanosine-12 and guanosine-14 are involved in the Cro-DNA interaction, while the three A·T base-pairs at the 3'- and 5'-termini play only a minor role in the binding. A model of the Cro-operator DNA complex has been constructed by docking helix-3 of the Cro protein in the major groove and it predicted specific hydrogen bonds between N7 of guanosines-12 and -14 and the side-chain of Lys-32 and Ser-28, respectively. The appearance of a new resonance in the temperature dependent proton detected heteronuclear multiple quantum coherence (HMQC) spectra of the Cro-DNA complex also demonstrates a specific interaction of Cro with guanosine-14 of the operator DNA.
23Na NMR Study of the Interaction
Between DNA and the Platinum (II) Compounds: Cis-DDP, Trans-DDP and TDP
Georges Mallet, Seddik Ansiss and Dan Vasilescu*
Laboratoire de Biophysique - GRECFO - Université de Nice-Sophia Antipolis
Parc Valrose, 06108 Nice Cedex 2, France
Abstract
The interaction of calf thymus DNA with cis-DDP, trans-DDP and TDP was studied by 23Na NMR in aqueous solutions at pH = 7.0, with Pt(II) compounds/DNA(P) (P=Phosphate) molar ratios r increasing from 0 till to 1P.
23Na NMR results are interpreted on one hand, with the help of a " two states model " with RF and RB relaxation rates, and, on the other hand, using the " entropy of fluctuations " concept developed by Lenk. We have established that, for the studied platinum compounds, the preference to interact with DNA phosphate sites -interpreted as a perturbation of the counterions environment- is in a decreasing order: TDP >> cis-DDP > trans-DDP. These results are discussed with regard to the interaction of DNA with the hard bication Mg++ and the soft bication Cu++.
Localization of Histone H1 Binding Sites Within the Nucleosome by UV-induced H1-DNA Crosslinking In Vivo
Sergey Belikov* and Vadim Karpov*
W. A. Engelhardt Institute of Molecular Biology,
Russian Academy of Sciences,
Laboratory of Structure and Function of Chromatin,
117984 Moscow, Russia
Abstract
In our previous paper (Belikov et al., (1993), Nucl. Acids Res., 21, 4796-4802) we had studied DNA-protein architecture of so-called Alu-repeats in D. melanogaster ribosomal non-transcribed spacer using DNase I genomic footprinting and UV-induced DNA-protein crosslinking. Our data indicated precise positioning of two non-histone proteins (rABP50 and rABP 70), histone octamer, and histone H1 within the sequences of Alu-repeats. The data on the histone H1 binding sites within Alu-repeat region was presented, but not discussed as the authors could not reach a consensus in its evaluation. Here, the authors use these data to present a model of placement of the linker histone(s) within nucleosome. Our model places one contact of the globular domain of linker histone in the major groove on the inside of DNA superhelix just within the end of the core particle (site +6.5) and the second, close to the dyad (site -1). C-terminal tail binds to the internucleosomal spacer and N-terminal tail interacts simultaneously with the adjacent gyres thus stabilizing DNA superhelix.
A Theoretical Perusal of
the Satellite DNA Curvature in Tenebrionid Beetles
Francisca Barceló1, Francisco
Gutiérrez1, Ignasi Barjau1
and José Portugal2*
1Departament de Biologia Fundamental i Ciencies
de la Salut,
Universitat de les Illes Balears,
Palma de Mallorca, Spain
2Departamento Biología Molecular y Celular,
Centro de Investigación y Desarrollo, CSIC,
J.Girona-Salgado, 18-26, 08034 Barcelona, Spain
Abstract
The curvature patterns of seven satellite DNAs taken from beetles belonging to the family Tenebrionidae (Coleoptera) were modelled utilising a number of computer programs that describe and plot the curvature profiles of DNA. The theoretical analysis agreed with the experimentally observed curvature of most of these satellite DNAs, and its absence in Tribolium freemani and Tenebrio obscurus satellite I. In many cases, the tenebrionid satellite DNAs lack periodically repeated runs of phased-A-tracts, yet they represent a clear example of cuved DNA. The macroscopic curvature of satellites from these closely related organisms confirmed that other sequence elements must be participating in the bending of these DNAs. Our modelling approaches are discussed, together with previous experimental results, in terms of the role played by DNA curvature in the organisation of satellite DNA and the tight compacting of heterochromatin.
The Base Contents of A, C,
G or U for the Three Codon Positions and the Total Coding Sequences Show
Positive Correlation
Ju Wang*
Chemical Engineering Research Center,
Tianjin University,
Tianjin 300072,China
Abstract
The distribution of the occurrence frequencies of each of the four bases at the first, second and third codon positions and in the total coding sequences is analyzed by a graphic method. It is shown that for the coding sequences of 90 species, A has its largest frequency at the second codon position and the smallest one at the third position. C and U have their least frequencies at the first codon position, while G has its largest frequency at the first codon position. By this method, we also find that for each base, there is positive correlation between every two frequencies of the base in the first, second, third and the total coding sequences for 90 species. For each of the four bases, the correlation between the frequencies at the third codon position and that in the total coding sequences is more prominent than others. A statistical method is used to give a precise description of the correlation for the frequencies of every base and it is found that the conclusions drawn by the graphic method are consistent with that got by the statistical method.
Melting of Cross-linked DNA.
III. Calculation of Differential Melting Curves.
Dmitri Y. Lando1*, Alexander S. Fridman1, Vladimir I. Krot2 and
Afanasii A. Akhrem1
1Institute of Bioorganic Chemistry,
Belarus National Academy of Sciences,
Zhodinskaya St. 5/2,
220141 Minsk, Belarus
2Byelorussian State University,
220050 Minsk, Belarus
Abstract
In our previous papers I and II (D. Y. Lando et al, J. Biomol. Struct.
Dynam. (1997) v. 15, N1, p.129-140, p. 141-150), two methods were developed
for calculation of melting curves of cross-linked DNA. One of them is based
on Poland's and another on the Fixman-Freire approach. In the present communication,
III, a new theoretical method is developed for computation of differential
melting curves of DNAs cross-linked by anticancer drugs and their inactive
analogs. As Poland's approach, the method allows study of the influence
of the loop entropy factor, d(n), on melting behavior (n is the length of
a loop in base pairs). However the method is much faster and requires computer
time that inherent for the most rapid Fixman-Freire calculation approach.
In contrast to the computation procedures described before in communications
I and II, the method is suitable for computation of differential melting
curves in the case of long DNA chains, arbitrary loop entropy factors of
melted regions and arbitrary degree of cross-linking including very low
values that occur in vivo after administration of antitumor drugs.
The method is also appropriate for DNAs without cross-links. The results
of calculation demonstrate that even very low degree of cross-linking alters
the DNA differential melting curve. Cross-linking also markedly strengthens
the influence of particular function d(n) upon melting behavior.
Molecular Structure of Two
Crystal Forms of Cyclic Triadenylic Acid at 1Å Resolution
Yi-Gui Gao1, Howard Robinson1, Yue Guan1, Yen-Chywan
Liaw1, Jacques H. van Boom2,
Gijs A. van der Marel2 and Andrew H.-J. Wang1*
1Department of Cell & Structural Biology,
University of Illinois at Urbana-Champaign,
Urbana, IL 61801 USA
2Leiden Institute of Chemistry,
Leiden State University,
Leiden, 2300 RA,The Netherlands
Abstract
The three dimensional structures of cyclic deoxytriadenylic acid, c-d(ApApAp),
from two different trigonal crystal forms (space groups P3 and R32)
have been determined by x-ray diffraction analysis at 1Å resolution.
Both structures were solved by direct methods and refined by anisotropic
least squares refinement to R-factors of 0.109 and 0.137 for the P3 and
R32 forms, respectively. In both crystal forms, each of the two independent
c-d(ApApAp) molecules sits on the crystallographic 3-fold axis. All four
independent c-d(ApApAp) molecules have similar backbone conformations. The
deoxyriboses are in the S-type pucker with pseudorotation angles
ranging from 156.7° to 168.6° and the bases have anti glycosyl
torsion angles (x falling in two ranges, one at -104.3° and the
other ranging from -141.0° to 143.8°). In the R32 form,
a hexahydrated cobalt(II) ion is found to coordinate through bridging water
molecules to N1, N3, and N7 atoms of three adjacent adenines and oxygen
atoms of phosphates. Comparison with other structures of cyclic oligonucleotides
indicates that the sugar adopts N-type pucker in cyclic dinucleotides
and S-type pucker in cyclic trinucleotides, regardless whether the
sugar is a ribose or a deoxyribose.
Molecular Modeling of HIV-1 Coreceptor CCR5 and Exploring of Conformational Space of its Extracellular Domain in Molecular Dynamics Simulation
Roman G. Efremov1*, François Legret2, Gérard Vergoten2,
André Capron3, Georges M. Bahr3 and Alexander S. Arseniev1
1M.M. Shemyakin and Yu.A. Ovchinnikov Institute of Bioorganic
Chemistry,
Russian Academy of Sciences,
Ul. Miklukho-Maklaya 16/10,
117871 GSP, Moscow V-437, Russia
2Université des Sciences et Technologies
de Lille,
Centre de Recherches et d'Etudes en Simulations et Modélisation Moléculaires
(CRESIMM),
Bâtiment C8, 59655 Villeneuve d'Ascq, Cedex, France
3Institut Pasteur de Lille, 1,
rue du Professeur Calmette, B.P. 245,
59019 Lille Cedex, France
Abstract
The chemokine receptor CCR5 functions as a major fusion coreceptor for macrophage-tropic human immunodeficiency virus entry into cell. Here we report a three-dimensional model of CCR5 built using molecular modeling approach. Because the virus binds to extracellular domain of the receptor, special attention was given to conformational flexibility, hydrogen bonding, and environmental polarity properties of this protein part. Such data were obtained in the result of molecular dynamics study of the extracellular domain. It was shown that during the simulation the extracellular segments form a compact globular domain with numerous long-range hydrogen bonds between them. First loop of the receptor stays quite rigid while N-terminal region and loops 2, 3 are rather flexible. A number of amino acid residues disposed in unfavourable environment and, therefore, potentially involved in binding of CCR5 to viral glycoproteins and chemokines, was delineated. Comparison of the results with available experimental data permits a proposal that such residues in loop-1 and N-terminal part of the receptor are important for HIV-1 entry, while those in loops 2 and 3 participate in ligand binding. Perspectives of rational alteration of virus-binding activity of CCR5 are discussed.
Delineation of Conformational
Preferences in Human Salivary Statherin by 1H, 31P NMR and CD Studies: Sequential Assignment and Structure-Function
Correlations
Gowda A. Naganagowda, Tarikere L. Gururaja and Michael J. Levine*
Department of Oral Biology and Dental Research Institute,
State University of New York at Buffalo,
Buffalo, NY 14214-3092
Abstract
Membrane-induced solution structure of human salivary statherin, a 43
amino acid residue acidic phosphoprotein, has been investigated by two-dimensional
proton nuclear magnetic resonance (2D 1H NMR) spectroscopy.
NMR assignments and structural analysis of this phosphoprotein was accomplished
by analyzing the pattern of sequential and medium range NOEs, alphaCH chemical
shift perturbations and deuterium exchange measurements of the amide proton
resonances. The NMR data revealed three distinct structural motifs in the
molecule: (1) an alpha-helical structure at the N-terminal domain comprising
Asp1-Tyr16, (2) a polyproline type II (PPII) conformation
predominantly occurring at the middle proline-rich domain spanning Gly19-Gln35, and (3) a 310-helical
structure at the C-terminal Pro36-Phe43
sequence. Presence of a few weak dalphaN(i,i+2)
NOEs suggests that N-terminus also possesses minor population of 310-helical conformation. Of the three secondary structural
elements, helical structure formed by the N-terminal residues, Asp1-Ile11 appears to be more rigid as
observed by the relatively very slow exchange of amide hydrogens of Glu5-Ile11. 31P NMR
experiments clearly indicated that N-terminal domain of statherin exists
mainly in disordered state in water whereas, upon addition of structure
stabilizing co-solvent, 2,2,2-trifluorethanol (TFE), it showed a strong
propensity for helical conformation. Calcium ion interaction studies suggested
that the disordered N-terminal region encompassing the two vicinal phosphoserines
is essential for the binding of calcium ions in vivo. Results from
the circular dichroism (CD) experiments were found to be consistent with
and complimentary to the NMR data and provided an evidence that non-aqueous
environment such as TFE, could induce the protein to fold into helical conformation.
The findings that the statherin possesses blended solvent sensitive secondary
structural elements and the requirement of non-structured N-terminal region
under aqueous environment in calcium ion interaction may be invaluable to
understand various physiological functions of statherin in the oral fluid.
The Influence of pH Alteration
and Pharmacological Modulators of Adenylate Cyclase System on Human Serum
Albumin Conformation
A.I. Luik*, Yu.N. Naboka, S.E. Mogilevich, T.O. Hushchaand N.I. Mischenko
Department of Biomedical Research,
Institute of Bioorganic Chemistry and Petrochemistry,
National Academy of Sciences of Ukraine,
Murmans'ka Str. 1, 253094 Kyiv 94, Ukraine
Laboratory MSC,
Chemical Department,
Catholic University of Leuven,
Celestijnenlaan 200 F, B -3001, Heverlee, Belgium
Abstract
The report describes the results of a study the effect of pH and binding of six physiologically active compounds (isoproterenol, yohimbine, theophylline, propranolol, clonidine and carbachol) on the molecular structure of human serum albumin (HSA) using dynamic light scattering. It was found that the albumin globule had the most compact configuration (Stokes diameter 59-62Å) at physiological pH 7.4. The changes in pH both increased to 8.0 and decreased to 5.4, resulting in the growth of globule size to 72-81Å. At acidic shift of pH an additional peak arose in the correlation spectra. This peak was caused by the light scattering on the structures with the Stokes diameters of 29 - 37Å, which conformed to the sizes of the albumin subdomains. The additional peak was not displayed at basic shift of pH.
The interaction with propranolol, clonidine and carbachol, which hinder adenylate cyclase (AdC) signaling system of a cell, initiated structural rearrangements similar to acidic transitions. Isoproterenol, yohimbine and theophylline, which activate AdC, caused the conformational changes of HSA similar to basic transitions.
Simulated Annealing for Alpha-Helical
Protein Folding: Searches in Vicinity of the "Molten Globule"
State
Igor V. Grigoriev*Ý, Alexandra
B. Rakhmaninova and Andrey A. Mironov
Research Institute for Genetics of Industrial Microorganisms,
1-st Dorozhny proezd, 1,
Moscow 113545, Russia
ÝCurrent address:
Chemistry Department,
University of California,
Berkeley, CA 94720
Abstract
A new model for simulation of protein folding of alpha-helical proteins with known secondary structure is proposed. We are dealing here with the analysis of alpha-helix packings rather than with a detailed atom structure of a whole protein. Starting from a random compact packing of the helices the search is focused on a vicinity of "molten globule" states of a protein. In contrast to the majority of the known approaches for estimation of a protein free energy we introduce a simplified potential of interactions with solvent and consider conformational energy of the loops in addition to mean-force potential. The model was applied to several globular alpha-helical proteins and demonstrated high prediction accuracy in comparison with other known models.
Resonance Raman and Infrared
Spectral Studies on Radical Anions of Model Photosynthetic Reaction Center
Quinones (Naphthoquinone Derivatives)
G. Balakrishnan1, A. Babaei2, A.J. McQuillan2 and S. Umapathy1*
1Department of Inorganic and Physical Chemistry,
Indian Institute of Science,
Bangalore 560 012, India
2Department of Chemistry,
University of Otago,
P.O. Box 56,
Dunedin, New Zealand
Abstract
Quinones play a vital role in the processes of electron transfer in bacterial
photosynthetic reaction centers. It is of interest to investigate photochemical
reactions involving quinones with a view to elucidate structure-function
relationships in biological processes. Resonance Raman and FTIR spectra
of electrochemically generated radical anions of 2-methyl-1,4-naphthoquinone,
and 2-methyl-3-phytyl-1,4-naphthoquinone, also known as Vitamin K3
and Vitamin K1, respectively, (model compound for QA in Rhodopseudomonas viridis, a bacterial photosynthetic
reaction center) have been reported. The same study has also been extended
to 1,4-naphthoquinone for comparison. The vibrational assignments were carried
out on the basis of comparison with our earlier time resolved resonance
Raman studies on photochemically generated radical anions of 1,4-naphthoquinone
and 2-methyl-1,4-naphthoquinone (Balakrishnan et al., J. Phys. Chem.,
100, (1996), 16472-16478). These in vitro results have been compared
with the reported vibrational spectral data under in vivo conditions.
Dependencies of Substitution
Steps Number on Hamming Distance Are Identical For One-parameter Discrete
Models of Both Direct and Parallel Genetic Diversity
Michael Yu. Shchelkanov1,2, Lev A. Soinov1, Vadim V. Zalunin1, Nicole
S. Starikov1,2, Andrei A. Natan1,Victor
B. Kireev1 and Edward V. Karamov2,3*
1Moscow Institute of
Physics and Technology,
Faculty of Physicochemical Biology, Department of Molecular Biophysics,
Dolgoprudny, Moscow region,
141700 Russia
2D. I. Ivanovsky Institute of Virology, Immunochemistry
Group,
Gamaley 16, Moscow,
123098 Russia
3Moscow State University,
Institute of Applied Research,
Laboratory of Virology,
Vorobyevy gory, Moscow,
119899 Russia
Abstract
With the help of previously introduced enumeration procedure (M.Yu. Shchelkanov, A.N. Yudin, A.V Antonov, N.S. Starikov, A.A. Vedenov, E.V. Karamov, J. Biomol. Struct. Dyn. 15, 217-229 (1997)) and probability distribution function for the enumeration after some substitution steps (M.Yu. Shchelkanov, L.A. Soinov, V.V. Zalunin, D.A. Gumennyi, A.N. Yudin, A.A. Natan, V.B. Kireev, E.V. Karamov, J. Biomol. Struct. Dyn. 15, N 4, (1998)) we have demonstrated that dependencies of replication acts number on Hamming distance are identical for one-parameter discrete models of both direct and parallel genetic diversity.
Stabilization of the Purine·Purine·Pyrimidine
DNA Base Triplets by Divalent Metal Cations
(Opinion Paper: Point)
Jiri Sponer*1,2, Michal Sabat*3, Jaroslav V. Burda4, Anne
M. Doody5, Jerzy Leszczynski6
and Pavel Hobza1
1J.Heyrovsky Institute of Physical Chemistry,
Academy of Sciences of the Czech Republic,
Dolejskova 3,
18223 Prague, Czech Republic
2Institute of Biophysics,
Academy of Sciences of the Czech Republic,
Krávopolská 135,
61265 Brno, Czech Republic
3Department of Chemistry,
University of Virginia,
Charlottesville, Virginia 22901, USA
4Department of Chemical Physics,
Faculty of Mathematics and Physics,
Charles University,
Ke Karlovu 3,
12116 Prague, Czech Republic
5Department of Biochemistry,
Cornell University,
Ithaca, NY 14853, USA
6Department of Chemistry,
Jackson State University,
Jackson, Mississippi 39217, USA
Purine·purine·pyrimidine (Pu·Pu·Py) DNA triplexes are formed by binding a purine-rich oligonucleotide to the purine strand of a DNA duplex. The third strand assumes an antiparallel orientation with respect to the purine strand of the duplex and the purine bases are associated by reverse Hoogsteen hydrogen bonding (1a-d). These DNA assemblies have recently been the subject of intense research because of their potential applications in the control of gene expression (2a-b). Affinity cleavage experiments (3) have indicated that the most stable Pu.PuPy triplexes consist of guanine.guanine-cytosine (G·G·C) and adenine.adenine-thymine (A·A·T) base triplets.
Stabilization of the Purine·Purine·Pyrimidine
DNA Base Triplets by Divalent Metal Cations
(Opinion Paper: Counterpoint)
Vladimir N. Potaman*1 and Valery N. Soyfer2
1Institute of Biosciences and Technology,
Texas A&M University,
2121 West Holcombe Blvd.,
Houston, TX 77030
2Department of Biology,
George Mason University,
4400 University Drive,
Fairfax, VA 22030
Understanding the stability of the Purine·Purine·Pyrimidine (Pu·Pu·Py) triplex has been a challenge for a number of years. It is generally accepted that the stabilization of this triplex requires multivalent cations. The stabilizing effects of polyamines and cationic peptides (1,2) can be understood in terms of electrostatic interactions between their distributed positive charges and anionic phosphate groups in the three polynucleotide chains. Less understood is the effect of divalent metal cations among which the transition metal cations are the most efficient triplex stabilizers (3). A strong stabilizing effect of transition metal cations might result from their tighter binding to phosphates due to shorter ionic radii (4). However, mechanisms other than electrostatic screening might be also involved. For example, a major difference in the triplex stabilization effects of the alkali-earth cation Mg2+ and the transition metal cation Mn2+ can hardly be explained by the little difference in their ionic radii (5).
Abstracts:
25th Anniversary Conference,
National Foundation for Cancer Research
Basic Science: Research for a Cure and Prevention
September 14-16, 1998
Georgetown Uiversity Conference Center,
Washington, DC