Biomedical subjects

P Auffinger

Publications and source records attributed to P Auffinger.

16 recordsLinked to original sources

PubMed2006

A molecular dynamics simulation study of an aminoglycoside/A-site RNA complex: conformational and hydration patterns.

Aminoglycoside antibiotics interfere with the translation mechanism by binding to the tRNA decoding site of the 16S ribosomal RNA. Crystallographic structures of aminoglycosides bound to A-site systems clarified many static aspects of RNA-ligand interactions. To gain some insight on the dynamic aspects of recognition phenomena, we conducted molecular dynamics simulations of the aminoglycoside paromomycin bound to a eubacterial ribosomal decoding A-site oligonucleotide. Results from 25 ns of simulation time revealed that: (i) the neamine part of the antibiotic represents the main anchor for binding, (ii) additional sugar rings provide limited and fragile contacts, (iii) long-resident water molecules present at the drug/RNA interface are involved in the recognition phenomena. The combination of MD simulations together with systematic structural information offers striking insights into the molecular recognition processes underlying RNA/aminoglycoside binding. Important methodological considerations related to the use of medium resolution starting structures and associated sampling problems are thoroughly discussed.

Aminoglycosides↗

PubMed2001

Water and ion binding around r(UpA)12 and d(TpA)12 oligomers--comparison with RNA and DNA (CpG)12 duplexes.

The structural and dynamic properties of the water and ion first coordination shell of the r(A-U) and d(A-T) base-pairs embedded within the r(UpA)12 and d(TpA)12 duplexes are described on the basis of two 2.4 ns molecular dynamics simulations performed in a neutralizing aqueous environment with 0.25 M added KCl. The results are compared to previous molecular dynamics simulations of the r(CpG)12 and d(CpG)12 structures performed under similar conditions. It can be concluded that: (i) RNA helices are more rigid than DNA helices of identical sequence, as reflected by the fact that RNA duplexes keep their initial A-form shape while DNA duplexes adopt more sequence-specific shapes. (ii) Around these base-pairs, the water molecules occupy 21 to 22 well-defined hydration sites, some of which are partially occupied by potassium ions. (iii) These hydration sites are occupied by an average of 21.9, 21.0, 20.1, and 19.8 solvent molecules (water and ions) around the r(G=C), r(A-U), d(G=C), and d(A-T) pairs, respectively. (iv) From a dynamic point of view, the stability of the hydration shell is the strongest for the r(G=C) pairs and the weakest for the d(A-T) pairs. (v) For RNA, the observed long-lived hydration patterns are essentially non-sequence dependent and involve water bridges located in the deep groove and linking OR atoms of adjacent phosphate groups. Maximum lifetimes are close to 400 ps. (vi) In contrast, for DNA, long-lived hydration patterns are sequence dependent and located in the minor groove. For d(CpG)12, water bridges linking the (G)N3 and (C)O2 with the O4' atoms of adjacent nucleotides with 400 ps maximum lifetimes are characterized while no such bridges are observed for d(TpA)12. (vii) Potassium ions are observed to bind preferentially to deep/major groove atoms at RpY steps, essentially d(GpC), r(GpC), and r(ApU), by forming ion-bridges between electronegative atoms of adjacent base-pairs. On average, about half an ion is observed per base-pair. Positive ion-binding determinants are related to the proximity of two or more electronegative atoms. Negative binding determinants are associated with the electrostatic and steric hindrance due to the proximity of electropositive amino groups and neutral methyl groups. Potassium ions form only transient contacts with phosphate groups.

AT Rich Sequence↗

PubMed2001

An extended structural signature for the tRNA anticodon loop.

Anticodon hairpins are structural motifs with contradictory functions. The recognition by aminoacyl synthetases implies extended interactions with the anticodon base triplet and thus, usually, an unfolding of the anticodon loop. The recognition by the ribosome and cognate interaction with a mRNA codon implies, on the other hand, the formation of a mini-helix with a canonical anticodon hairpin structure as observed by crystallography and NMR. To be able to understand the various properties of this motif, a precise description of its structural conservation is required. Here, on the basis of phylogenetic, structural, and molecular dynamics data, we discuss a conserved interaction established between the ribose of the U33 and the base at position 35, either a purine or a pyrimidine. This interaction involves the hydrogen bonding donor or acceptor potential of the hydroxyl group of U33 and has to be integrated in an extended definition of the anticodon hairpin. The extended structural signature provides also an explanation for the role played by pseudouridines at position 35.

Anticodon↗

PubMed2000

Water and ion binding around RNA and DNA (C,G) oligomers.

The dynamics, hydration, and ion-binding features of two duplexes, the A(r(CG)(12)) and the B(d(CG)(12)), in a neutralizing aqueous environment with 0.25 M added KCl have been investigated by molecular dynamics (MD) simulations. The regular repeats of the same C=G base-pair motif have been exploited as a statistical alternative to long MD simulations in order to extend the sampling of the conformational space. The trajectories demonstrate the larger flexibility of DNA compared to RNA helices. This flexibility results in less well defined hydration patterns around the DNA than around the RNA backbone atoms. Yet, 22 hydration sites are clearly characterized around both nucleic acid structures. With additional results from MD simulations, the following hydration scale for C=G pairs can be deduced: A-DNA<RNA (+3 H(2)O) and B-DNA<RNA (+2 H(2)O). The calculated residence times of water molecules in the first hydration shell of the helices range from 0.5 to 1 ns, in good agreement with available experimental data. Such water molecules are essentially found in the vicinity of the phosphate groups and in the DNA minor groove. The calculated number of ions that break into the first hydration shell of the nucleic acids is close to 0.5 per base-pair for both RNA and DNA. These ions form contacts essentially with the oxygen atoms of the phosphate groups and with the guanine N7 and O6 atoms; they display residence times in the deep/major groove approaching 500 ps. Further, a significant sequence-dependent effect on ion binding has been noted. Despite slight structural differences, K(+) binds essentially to GpC and not to CpG steps. These results may be of importance for understanding various sequence-dependent binding affinities. Additionally, the data help to rationalize the experimentally observed differences in gel electrophoretic mobility between RNA and DNA as due to the difference in hydration (two water molecules in favor of RNA) rather than to strong ion-binding features, which are largely similar for both nucleic acid structures.

Base Pairing↗

PubMed1999

Singly and bifurcated hydrogen-bonded base-pairs in tRNA anticodon hairpins and ribozymes.

The tRNA anticodon loops always comprise seven nucleotides and is involved in many recognition processes with proteins and RNA fragments. We have investigated the nature and the possible interactions between the first (32) and last (38) residues of the loop on the basis of the available sequences and crystal structures. The data demonstrate the conservation of a bifurcated hydrogen bond interaction between residues 32 and 38, located at the stem/loop junction. This interaction leads to the formation of a non-canonical base-pair which is preserved in the known crystal structures of tRNA/synthetase complexes. Among the tRNA and tDNA sequences, 93 % of the 32.38 oppositions can be assigned to two families of isosteric base-pairs, one with a large (86 %) and the other with a much smaller (7 %) population. The remainder (7 %) of the oppositions have been assigned to a third family due to the lack of evidence for assigning them into the first two sets. In all families, the Y32.R38 base-pairs are not isosteric upon reversal (like the sheared G.A or wobble G.U pairs), explaining the strong conservation of a pyrimidine at position 32. Thus, the 32.38 interaction extends the sequence signature of the anticodon loop beyond the conserved U-turn at position 33 and the usually modified purine at position 37. A comparison with other loops containing both a singly hydrogen-bonded base-pair and a U-turn suggests that the 32.38 pair could be involved in the formation of a base triple with a residue in a ribosomal RNA component. It is also observed that two crystal structures of ribozymes (hammerhead and leadzyme) present similar base-pairs at the cleavage site.

Amino Acyl-tRNA Synthetases↗

PubMed1999

Molecular dynamics simulations of solvated yeast tRNA(Asp).

Transfer RNA molecules are involved in a variety of biological processes, implying complex recognition events with proteins and other RNAs. From a structural point of view, tRNAs constitute a reference system for studying RNA folding and architecture. A deeper understanding of their structural and functional properties will derive from our ability to model accurately their dynamical behavior. We present the first dynamical model of a fully neutralized and solvated tRNA molecule over a 500-ps time scale. Starting from the crystallographic structure of yeast tRNA(Asp), the 75-nucleotide molecule was modeled with 8055 water molecules and 74 NH4+ counterions, using the AMBER4.1 program and the particle mesh Ewald (PME) method for the treatment of long-range electrostatic interactions. The calculations led to a dynamically stable model of the tRNA molecule. During the simulation, all secondary and tertiary base pairs are maintained while a certain lability of base triples in the tRNA core is observed. This lability was interpreted as resulting from intrinsic factors associated with the "weaker" hydrogen bonding patterns seen in these base triples and from an altered ionic environment of the tRNA molecule. Calculated thermal factors are used to compare the dynamics of the tRNA in solution and in the crystal. The present molecular dynamics simulation of a complex and highly charged nucleic acid molecule attests to the fact that simulation methods are now able to investigate not only the dynamics of proteins, but also that of large RNA molecules. Thus they also provide a basis for further investigations on the structural and functional effects of chemical and posttranscriptionally modified nucleotides as well as on ionic environmental effects.

Base Pairing↗

PubMed1998

Molecular dynamics investigations of hammerhead ribozyme RNA.

The hammerhead ribozyme, a small catalytic RNA molecule, cleaves, in the presence of magnesium ions, a specific phosphodiester bond within its own backbone, leading to 23-cyclic phosphate and 5-OH extremities. In order to study the dynamical flexibility of the hammerhead RNA, we performed molecular dynamics simulations of the solvated crystal structure of an active hammerhead ribozyme, obtained after flash-freezing crystals soaked with magnesium. Because of a careful equilibration protocol and the use of the Ewald summation in calculating the electrostatic interactions, the RNA structure remained close to the crystal structure, as attested by a root-mean-square deviation below 2.5 A after 750 ps of simulation. All Watson-Crick base pairs were intact at the end of the simulations. The tertiary interactions, such as the sheared G.A pairs and the U-turn, important for the stabilisation of the three-dimensional RNA fold, were also retained. The results demonstrate that molecular dynamics simulations can be successfully used to investigate the dynamical behaviour of a ribozyme, thus, opening a road to study the role of transient structural changes involved in ribozyme catalysis.

Base Pairing↗

PubMed1998

Simulations of the molecular dynamics of nucleic acids.

The growing amount of high quality molecular dynamics simulations generated using the latest methodological developments and force fields has led to a sharper understanding of the forces underlying the dynamics of biomolecular systems, as well as to stimulating insights into the structure and catalysis of nucleic acids. It is now clear that inclusion of long-range electrostatic interactions and of the aqueous and ionic environment is necessary for producing realistic and accurate simulations. Yet, many papers hint at a force field and protocol dependence of the results and thus contain the seeds for the future improvements that will be necessary for deepening our understanding of recognition phenomena and folding of nucleic acids.

Crystallization↗

PubMed1998

Hydration of RNA base pairs.

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.

Base Pair Mismatch↗

PubMed1997

Rules governing the orientation of the 2'-hydroxyl group in RNA.

Molecular dynamics simulations reveal that, in C3'-endo sugar puckers, only three orientations are accessible to the 2'-hydroxyl groups distinctive of RNA molecules: towards (i) the O3', (ii) the O4' of the same sugar, and (iii) the shallow groove base atoms. In the rarer C2'-endo sugar puckers, orientations towards the O3' atom of the same sugar are strongly favoured. Surprisingly, in helical regions, the frequently suggested intra-strand O2'-H(n)...O4'(n+1) interaction is not found. This observation led to the detection of an axial C-H...O interaction between the C2'-H2'(n) group and the O4'(n+1) atom contributing to the stabilization of RNA helical regions. Subsequent analysis of crystallographic structures of both RNA and A-DNA helices fully supports this finding. Specific hydration patterns are also thought to play a significant role in the stabilization of RNA structures. In the shallow groove of RNA, known as a favourable RNA or protein-binding region, three well-defined hydration sites are located around the O2' atom. These hydration sites, occupied by water molecules exchanging with the bulk, constitute, after dehydration, anchor points for specific interactions between RNA and nucleic acids, proteins or drugs. Therefore, the fact that the 2'-hydroxyl group is not monopolised by axial stabilization, together with its water-like behaviour, facilitates complex formation involving RNA helical regions.

Carbohydrate Conformation↗

PubMed1997

Evidence for a hydroxide ion bridging two magnesium ions at the active site of the hammerhead ribozyme.

In the presence of magnesium ions, cleavage by the hammerhead ribozyme RNA at a specific residue leads to 2'3'-cyclic phosphate and 5'-OH extremities. In the cleavage reaction an activated ribose 2'-hydroxyl group attacks its attached 3'-phosphate. Molecular dynamics simulations of the crystal structure of the hammerhead ribozyme, obtained after flash-freezing of crystals under conditions where the ribozyme is active, provide evidence that a mu-bridging OH-ion is located between two Mg2+ions close to the cleavable phosphate. Constrained simulations show further that a flip from the C3'- endo to the C2'- endo conformation of the ribose at the cleavable phosphate brings the 2'-hydroxyl in proximity to both the attacked phosphorous atom and the mu-bridging OH-ion. Thus, the simulations lead to a detailed new insight into the mechanism of hammerhead ribozyme cleavage where a mu-hydroxo bridged magnesium cluster, located on the deep groove side, provides an OH-ion that is able to activate the 2'-hydroxyl nucleophile after a minor and localized conformational change in the RNA.

Binding Sites↗

PubMed1997

RNA hydration: three nanoseconds of multiple molecular dynamics simulations of the solvated tRNA(Asp) anticodon hairpin.

The hydration of the tRNA(Asp) anticodon hairpin was investigated through the analysis of six 500 ps multiple molecular dynamics (MMD) trajectories generated by using the particle mesh Ewald method for the treatment of the long-range electrostatic interactions. Although similar in their dynamical characteristics, these six trajectories display different local hydration patterns reflecting the landscape of the "theoretical" conformational space being explored. The statistical view gained through the MMD strategy allowed us to characterize the hydration patterns around important RNA structural motifs such as a G-U base-pair, the anticodon U-turn, and two modified bases: pseudouridine and 1-methylguanine. The binding of ammonium counterions to the hairpin has also been investigated. No long-lived hydrogen bond between water and a 2'-hydroxyl has been observed. Water molecules with long-residence times are found bridging adjacent pro-Rp phosphate atoms. The conformation of the pseudouridine is stiffened by a water-mediated base-backbone interaction and the 1-methylguanine is additionally stabilized by long-lived hydration patterns. Such long-lived hydration patterns are essential to ensure the structural integrity of this hairpin motif. Consequently, our simulations confirm the conclusion reached from an analysis of X-ray crystal structures according to which water molecules form an integral part of nucleic acid structure. The fact that the same conclusion is reached from a static and a dynamic point of view suggests that RNA and water together constitute the biologically relevant functional entity.

Anticodon↗

PubMed1996

H-bond stability in the tRNA(Asp) anticodon hairpin: 3 ns of multiple molecular dynamics simulations.

Multiple molecular dynamics trajectories of the solvated and neutralized 17-residue tRNA(Asp) anticodon hairpin were generated for a total of 3 ns. Explicit treatment of all long-ranged electrostatic interactions by the particle mesh Ewald algorithm, as implemented in the AMBER MD software package, effected a degree of structural stabilization not previously achieved by use of a long 16-A solvent interaction truncation scheme. The increased stability of this multiple molecular dynamics set was appropriate for an in-depth analysis of the six 500-ps-long trajectories and allowed the characterization of a number of key structural interactions. The dynamical behavior of the standard Watson-Crick base pairs, the noncanonical G30-U40 "wobble" base pair, and the psi 32-C38 pseudo-base pair is presented as well as that of two C--H... O hydrogen bonds found to contribute to the array of tertiary interactions that stabilize the seven-nucleotide native loop conformation. The least mobile residue in the loop is U33, which forms the U-turn motif and which participates in several hydrogen-bonding interactions, whereas the most mobile residue is the apical residue G34 at the wobble position, a factor undoubtedly important in its biological function. The set of multiple molecular dynamics trajectories obtained does not converge on a 500-ps time scale to a unique dynamical model but instead describes an ensemble of structural microstates accessible to the system under the present simulation protocol, which is the result of local structural heterogeneity rather than of global conformational changes.

Anticodon↗

PubMed1996

Calculations of nucleic acid conformations.

The present computational power and sophistication of theoretical approaches to nucleic acid structural investigation are sufficient for the realization of static and dynamic models that correlate accurately with current crystallographic, NMR and solution-probing structural data, and consequently are able to provide valuable insights and predictions for a variety of nucleic acid conformational families. In molecular dynamics simulations, the year 1995 was marked by the foray of fast Ewald methods, an accomplishment resulting from several years' work in the search for an adequate treatment of the electrostatic long-range forces so primordial in nucleic acid behavior. In very large systems, and particularly in the RNA-folding field, techniques originating from artificial intelligence research, like constraint satisfaction programming or genetic algorithms, have established their utility and potential.

DNA↗

PubMed1996

Hydration of C-H groups in tRNA.

Molecular dynamic (MD) simulations of the anticodon hairpin of tRNA(Asp) and of the full tRNA, both in a solvent bath with neutralizing NH4+ counter-ions, have been produced with the particle mesh ewald (PME) method and the multiple molecular dynamics (MMD) strategy. The latter consists of generating uncorrelated trajectories starting from the same initial configuration but with a slightly perturbed initial velocity distribution. The 3 ns (six uncorrelated 500 ps MD trajectories) MMD set of the 17 nucleotide anticodon fragment and the single 500 ps trajectory of the 75 nucleotide tRNA were analyzed with the aim of characterizing long lived C-H...Ow interactions for the two main nucleic acid base and ribose C-H bond types. Some C-H sites present very long residence lifetimes for water molecules, especially those around the ribose H(3') and the pyrimidine H(5) atoms. The C(3')-H(3')...Ow contacts occur concurrently with the strong hydration of the anionic phosphate oxygen atoms and especially with the water bridges linking successive phosphate groups along the polynucleotide chain. Therefore, these contacts are of opportunistic character and result from the geometries of the covalent structure and adjacent interactions. On the other hand, the pyrimidine H(5) atoms display a hydrophilic character with interaction geometries indicating that water contacts in which they are involved should be considered as bona fide hydrogen bonds.

Base Sequence↗

PubMedDate not supplied

RNA solvation: a molecular dynamics simulation perspective.

With the availability of accurate methods to treat the electrostatic long-range interactions, molecular dynamics simulations have resulted in refined dynamical models of the structure of the hydration shell around RNA motifs. The models reviewed here range from basic Watson-Crick to more specific noncanonical base pairs, from "simple" double helices to RNA molecules displaying more complex tertiary folds, and from DNA/RNA hybrid double helices to RNA hybrids formed with a chemically modified strand.

Base Pairing↗