bluuebunny/crossref_metadata_2025 huggingface dataset:
Helix‐Coil transition in polypeptides: A microscopical approach
Authors: Ananikyan N. S. , Hajryan Sh. A. , Mamasakhlisov E. Sh. , Morozov V. F.
Published: December 2004

Analogous with the Potts model that describes the helix‐coil transition in the isolated polypeptide chain (a Hamiltonian model allowing for the energy U of hydrogen bond formation) the number Q of conformational states of a repeating unit of the chain and the topology of Δ = 3 hydrogen bond formation (the hydrogen bond fixing three pairs of φψ chain rotations) has been constructed and the corresponding transfer‐matrix has been obtained. In the thermodynamical limit, the partition function is expressed through the principal root of the cubic equation. The degree of helicity, the transition point and range, the correlation length, the number of junctions between the helical and coiling sections as well as the mean length of helical and coiling sections are calculated. Empirically introduced parameters of the Zimm–Bragg theory, constants of hydrogen bond formation s, and the cooperativity parameter σ as functions of microscopic parameters U, Q, and Δ are obtained by direct calculations. The behavior of this model was investigated at other topologies of the hydrogen‐bond closing Δ = 2 and Δ = 4, and it was suggested that the actual polypeptide chain (Δ = 3) provides the optimum correlation of helical structure of the order of globule dimensions. An expression was obtained for the maximum correlation length of the order ξ ∼ Q(Δ‐1)/2. For a System with solvent competing for the formation of hydrogen bonds with peptide groups a Hamiltonian model was constructed that took account of the energy E of the formation of hydrogen bond with the solvent and the number q of orientations of a solvent molecule about the peptide groups. It is shown that by the redefinition of the temperature parameter, the model with solvent reduces to the model of an isolated chain. Aside from the definition relationship that exists between the parameters of the theory U < 2E < Uq and the ordinary helix‐coil transitions (“melting”), the model also describes the transition from the coiling state to the helical one (“arrangement”) under heating. The change in temperature and transition range with solvent parameters was discussed and it was shown, that despite the difference in ΔT for the given polypeptide chain (Q = constant) with different solvent parameters, at “melting” and “arrangement,” the transition occurred at the same correlation length (the same cooperativity).
Bicyclic peptides: Solid state conformation of cyclo(Glu‐Leu‐Pro‐Gly‐Lys‐Leu‐Pro‐Gly)cyclo(1γ‐5ϵ)Gly
Authors: Di Blasio Benedetto , Benedetti Ettore , Pavone Vincenzo , Pedone Carlo , Saviano Michele , Zanotti Giancarlo , Blout Elkan R.
Published: December 2004

The solid state conformational analysis of the ionophoric homodetic bicyclic cyclo(Glu‐Leu‐Pro‐Gly‐Lys‐Leu‐Pro‐Gly)cyclo(1γ‐5ϵ)Gly (BCP3) has been carried out by x‐ray diffraction. It crystallizes with 4.5 molecules of water per peptide molecule in the monoclinic system, space group P21, with a = 11.425 Å, b = 16.616 Å, c = 13.931 Å, β = 109.24°, and Z = 2. The structure has been determined by direct methods and refined to an R factor of 0.061 for 2448 observed reflections. The structure characterized by all trans peptide bonds is stabilized by three intramolecular hydrogen bonds: a type II β‐turn, a mixed type I‐type III β‐turn, and a pseudo γ‐turn, which involves the side chain CO and the main‐chain NH groups of the Glu1 residue. The resulting globular molecule presents a rather hydrophilic surface with most of the CO groups available to hydration of the solvent molecules, which are linked through hydrogen bonds of the NH … O or OH … O types in a complicated H‐bonding scheme. The conformation observed in the solid state is rather different from the conformation proposed in solution for both the free and the Ca2+‐complexed BCP3 molecule.
A study of model energetics and conformational properties of polynucleotide triplexes
Authors: Van Vlijmen Herman W. Th. , Rameé Graciela L. , Pettitt B. Montgomery
Published: December 2004

The formation of triple‐stranded nucleic acid helices is studied by molecular mechanics and molecular dynamics calculations. Using standard TAT and CGG homopolymers, single, triple, and quintuple molecular replacements are made. Some of these replacements are expected to form Hoogsteen bonds and some are not. While the electrostatic and total energetic differences for base triplet mismatches were dependent on the electrostatic model chosen, clear trends in the local geometric distortions were apparent. Relationships between these model‐built strand geometries and chemical probe experiments are discussed.
Hydrodynamic study of flexibility in immunoglobulin igG1 using Brownian dynamics and the Monte Carlo simulations of a simple model
Authors: Díaz F. G. , Iniesta A. , De La Torre J. García
Published: December 2004

A simple bead model is proposed for the antibody molecule immunoglobulin IgG1. The partial flexibility of the hinge is represented by a quadratic potential associated to the angles between arms. Conformational and hydrodynamic properties are calculated using Monte Carlo (rigid‐body) and Brownian dynamics simulations. Comparison of experimental and calculated values for some overall properties allows the assignment of dimensions and other model parameters. The Brownian dynamics technique is used next to simulate a rotational correlation function that is comparable with the decay of fluorescence emission anisotropy. This is done with varying flexibility at the hinge. The longest relaxation time shows a threefold decrease when going from the rigid Y‐shaped conformation to the completely flexible case. The calculations are in good agreement with the decay times observed for IgG1. A flexibility analysis of the latter indicates that a variability of ±55° (standard deviation) in the angle between the Fab arms.
The effect of conformation on the CD of interacting helices: A theoretical study of tropomyosin
Authors: Cooper Thomas M. , Woody Robert W.
Published: December 2004

A recent report [M. E. Holtzer, et al. (1988) Biophysics Journal, 53, 96a] of the anomalous CD spectum of the tropomyosin (TM) fragment 11TM127 motivated us to model the system as two 21‐residue α‐helices distorted to a coiled‐coil conformation. We used strong‐coupling exciton theory to model the optical properties of the system. Two backbone amide excited states (nπ* and ππ*) were considered, as well as four excited states (Lb, La, Bb, Ba) for the phenolic side chain. We calculated the effect of superhelix formation on the backbone CD spectrum. The decrease in molar ellipticity of the α‐helix parallel‐polarized transition at 208 nm was found to be a simple function of superhelix tilt angle. We then modeled a coiled coil (radius = 5.5 Å, pitch = −140 Å) with one aromatic ring per superhelix. Steric interactions between aromatic side chains in a coiled coil were calculated as a function of side‐chain conformation and heptet position. Steric interactions between phenolic rings will be significant for heptet positions a and d, but not for positions b, c, e, f, or g. We calculated the phenolic Lb transition rotational strength as a function of position within the heptet repeats, and of all possible side‐chain dihedral angles, X1 and X2. When tyrosines were placed at heptet positions b, c, e, f, or g, the rotational‐strength surface was nearly identical to that of a single tyrosine in an undistorted helix. In contrast, the rotational‐strength surface for tyrosines in heptet positions a or d showed substantial intertyrosine coupling components. The rotational‐strength surfaces for the three types of heptet positions (position a, position d, and the others) allowed an interpretation of the aromatic CD spectra of TM and its fragments. It was predicted that the three types of heptet positions will be spectroscopically distinguishable.
A 23Na‐nmr study of sodium–DNA interactions in concentrated DNA solutions at low‐supporting electrolyte concentration
Authors: Strzelecka Teresa E. , Rill Randolph L.
Published: December 2004

Aqueous solutions of DNA fragments with a contour length (500 Å) near the persistence length at DNA concentrations ranging from 10 to 290 mg/mL solvent and a constant supporting electrolyte concentration of 0.01 M (predominantly NaCl) were examined by 23Na‐nmr spectroscopy at temperatures of 20, 40, and 60°C. Over the higher portion of this concentration range (> 100 mg/ml) the DNA solutions undergo a complex series of transitions between different anisotropic, liquid crystalline phases (T. E. Strzelecka and R. L. Rill, Biopolymers, in press). Counterions in solutions of strong polyelectrolytes are usually described in terms of a two‐state model as free or “bound” (influenced by the electrostatic field of the polyanion). The longitudinal relaxation rate (R1 = 1/T1) at all DNA concentrations decreased with increasing temperature, demonstrating fast exchange between free and bound counterions. R1, increased nearly linearly with increasing DNA phosphate/sodium ratio in the isotropic domain until the onset of anisotropic phase formation, in agreement with similar nmr studies conducted at low DNA concentrations. The value of R1,b = 194 ± 7 Hz obtained for the isotropic phase from 10 to 100 mg DNA/mL at 20°C was in agreement with values reported previously. A nonlinear increase in R1 with DNA concentration was observed upon onset of anisotropic phase formation, indicating an increase in the product of the fraction of bond ions times their relaxation rate (r·R1,b).

The spectral lineshape of all isotropic samples was Lorentzian. Spectra of anisotropic samples exhibited low magnitude quadrupole splitting of ≤400 Hz correlated with appearance of a cholesteric phase with pitch ≈ 2 μm. The magnitude of the quadrupole splitting decreased with increasing DNA concentration at low temperatures and increased with concentration at high temperature. At all concentrations the quadrupole splitting decreased then increased with temperature. These temperature‐ and concentration‐dependent changes in quadrupole splitting are consistent with an angle between the DNA helix axis and the principle component (VZZ) of the local electric field gradient tensor near the “magic angle” of 54.7°.
Internal dynamics of tRNAPhe studied by depolarized dynamic light scattering
Authors: Patkowski A. , Eimer W. , Dorfmüller Th.
Published: December 2004

The collective internal dynamics of transfer RNAphe from brewer's yeast in solution was studied by depolarized dynamic light scattering (DDLS). Within the melting region of tRNA the depolarized spectra consist of two Lorentzian, where the narrow (slow) component describes the overall rotation of the macromolecule. The broad component is attributed to the collective reorientation of the bases within the biopolymer. At high temperature only this relaxation process is observed in the spectrum. The viscosity dependence of the collective internal relaxation process is described by the Stokes–Einstein–Debye equation for rotational diffusion. Estimates of the internal orientational pair correlation factor from the integral depolarized intensities of tRNAPhe solutions indicates that the observed dynamics correspond to the collective reorientation of approximately 5 bases. A comparison of the results presented with DDLS studies on the aggregation of the mononucleotide guanosine‐5′‐monophosphate confirms this result. For a further characterization of the relaxation process we studied the effect of hydrostatic pressure (1–1000 bar) on the depolarized spectra of tRNA. While other spectroscopic methods like nmr, fluorescence polarization anisotropy decay, or ESR give information about the very local motion of a single base within the DNA or RNA, this study shows that by DDLS one can characterize collective internal motions of macromolecules.
Random coil scission rates determined by time‐dependent total intensity light scattering: Hyaluronate depolymerization by hyaluronidase
Authors: Reed Wayne F. , Reed Christopher E. , Byers Larry D.
Published: December 2004

A recently developed theory of the light scattering by random coils undergoing random scission is applied to the digestion of hyaluronate by hyaluronidase. The time dependence of the scattered light from solutions undergoing digestion was monitored. Working at a high angle with high molecular weight hyaluronate allowed the use of a powerful approximation for determining initial velocities and the Henri–Michaelis–Menten coefficients, without explicit knowledge of the hyaluronate molecular weight, radius of gyration, second virial coefficient, or polydispersity. Effects due to a molecular weight dependent second virial coefficient and to non‐Gaussian behavior are briefly considered. Assays were performed over nearly two orders of magnitude in substrate concentration. The initial velocities are compared with those obtained by a standard reducing sugar assay, which was performed on identical samples. The main advantages of the light scattering assay procedure over the more traditional assays are that many relatively high‐precision data points can be quickly and automatically collected with simple apparatus, and that the technique is most sensitive for the initial period of digestion, where the other assays are least sensitive. The shapes of the scattering curves also provide evidence that hyaluronate in these solutions is not a stable double strand and that the hyaluronidase cleaves bonds randomly. The curves also indicate that enzyme deactivation occurs, which accounts for the lower velocities yielded by the slower reductimetric assay, which is measured over longer initial periods.
Restrained and unrestrained molecular dynamics simulations in the NVT ensemble of alamethicin
Authors: Fraternali F.
Published: December 2004

Molecular dynamics simulations on the transmembrane antibiotic peptide alamethicin have been performed in the NVT ensamble (i.e., the number of particles N, the volume V, and the temperature T of the system are kept constant). Results on the structure and conformational flexibility of this molecule are discussed and compared with previous experimental CD, x‐ray, nmr data and theoretical computations on fragments analogues.

An extensive study of structural and dynamic properties from H‐bonding pattern analysis is presented. Evidences for a largely α‐helix structure with some extent of freedom in the C‐terminal region are found.

Further, a partition of the molecule into three regions on the base of structural features and dynamic behavior has been proposed, and the correlation among the motions of the three regions is described.
Solvent‐ and salt‐induced coil–helix transition of alkali metal salts of poly(L‐glutamic acid) in aqueous organic solvents
Authors: Satoh Mitsuru , Fujii Yoshitake , Kato Fusaaki , Komiyama Jiro
Published: December 2004

The coil–helix transition has been studied for alkali metal salts of poly (L‐glutamic acid) (PLG), i.e., PLGLi, ‐Na, ‐K, and ‐Cs, in aqueous organic solvent systems. Dependence of the transition on the solvent composition has been qualitatively discussed in terms of the solvent dielectric constant D, Gutmann's acceptor number AN, and water activity aw. The helix formation induced by addition of alkali chlorides has also been studied. The sharpness of the transition has been interpreted as a measure of reduction of electrostatic energy of helical PLG through contact ion‐pair formation between a counterion and carboxyl anion.
Interaction of guanylic acid with the Mg(II), Ca(II), Sr(II), and Ba(II) ions in the crystalline solid and aqueous solution: Evidence for the ribose C2′‐endo/anti and C3′‐endo/anti conformational changes
Authors: Tajmir‐Riahi Heidar‐Ali
Published: December 2004

The interaction of guanosine‐5′‐monophosphoric acid (H2‐GMP) with the alkaline earth metal ions has been studied in aqueous solution at neutral pH. The crystalline salts of the type Mg‐GMP·5H2O, Ca‐GMP·6H2O, Sr‐GMP·7H2O, and Ba‐GMP·7H2O were isolated and characterized by Fourier transform ir, 1H‐nmr and x‐ray powder diffraction measurements. Two types of macrochelate complexes have been identified: (a) The direct metal–base and indirect metal–phosphate bindings (inner and outer sphere interaction) for the Mg(II), Ca(II), and Sr(II), ions; and (b) the indirect metal–base and direct metal–phosphate bindings (outer and inner sphere interaction) for the Ba(II) ion. In aqueous solution, an equilibrium exists between the base–metal–H2O…PO3 and base … H2O–M–PO3 interactions. The ribose moiety shows C3′‐endo/anti conformation in the free acid; C2′‐endo/anti in the Na2‐GMP salt; C3′‐endo/anti in the Mg(II)‐, Ca(II)‐, and Sr(II)‐GMP salts; and C2′‐endo/anti, in the Ba(II)‐GMP salt.
Transient electric birefringence study of highly dilute agarose solutions
Authors: Dormoy Y. , Candau S.
Published: December 2004

In order to characterize the first step of agarose gelation, highly dilute solutions (2·10−3 to 0.5 g/L) have been studied by means of the transient electric birefringence technique. The field‐free decay curves of the birefringence are well described by a stretched‐exponential B(t) ≈ exp(−t/τ)β; the value of the exponent β is close to 0.5 whatever the agarose concentration. The suspended particles observed by electron microscopy present a rod‐like shape with a constant diameter (∼50 Å), without any branching; they are polydisperse with a distribution of lengths approximately exponential. The mean length of these fibers, deduced from their mean rotational diffusion coefficient, is proportional to the 0.37 power of the agarose concentration in the solution. Furthermore, these particles possess a strong permanent electrical dipole confirming the side‐to‐side arrangement of helices into bundles; this dipole is roughly proportional to the particle length, indicating a self‐similarity in the unidirectional growth of the agarose fibers, even when approaching the gelling concentration.
Study of the “molten globule” intermediate state in protein folding by a hydrophobic fluorescent probe
Authors: Semisotnov G. V. , Rodionova N. A. , Razgulyaev O. I. , Uversky V. N. , Gripas' A. F. , Gilmanshin R. I.
Published: December 2004

Binding of the hydrophobia fluorescent probe, 1‐anilino‐naphthalene‐8‐sulfonate (ANS), to synthetic polypeptides and proteins with a different structural organization has been studied. It has been shown that ANS has a much stronger affinity to the protein “molten globule” state, with a pronounced secondary structure and compactness, but without a tightly packed tertiary structure as compared with its affinity to the native and coil‐like proteins, or to coil‐like, α‐helical, or β‐structural hydrophilic homopolypeptides.

The possibility of using ANS for the study of equilibrium and kinetic molten globule intermediates is demonstrated, with carbonic anhydrase, β‐lactamase, and α‐lactalbumin as examples.
Preferred conformation of the terminally blocked (Aib)10 homo‐oligopeptide: A long, regular 310‐helix
Authors: Toniolo Claudio , Crisma Marco , Bonora Gian Maria , Benedetti Ettore , Dl Blasio Benedetto , Pavone Vincenzo , Pedone Carlo , Santini Antonello
Published: December 2004

The decapeptide pBrBz‐ (Aib)10‐OtBu, synthesized by the 5(4H)‐oxazolone method, crystallizes in the monoclinic space group C2/c with a = 43.901(2), b = 9.289(2), and c = 34.746(3) A; β = 114.69(3)°; and Z = 8. The crystals contain one molecule of water associated with each peptide. The structure has been solved by the Patterson method and refined to an R value of 0.073 for 6819 observed reflections. The peptide adopts a regular 310‐helical structure stabilized by eight NH …︁ OC intramolecular 1 ← 4 (or C10) H bonds. This study has allowed us to characterize this important peptide secondary structure in great detail. The crystal‐state conformation agrees well with proposals made on the basis of an ir absorption and 1H‐nmr study in solution.
A molecular dynamics study of the effect of G · T mispairs on the conformation of DNA in solution
Authors: Shibata Masayuki , Zielinski Theresa Julia , Rein Robert
Published: December 2004

The effect of G · T mispair incorporation into a double‐helical environment was examined by molecular dynamics simulation. The 60‐ps simulations performed on the two hexanucleotide duplexes d (G3C3)2 and d (G3TC2)2 included 10 Na+ counterions and first hydration shell waters. The resulting backbone torsional angle trajectories were analyzed to select time spans representative of conformational domains. The average backbone angles and helical parameters of the last time span for both duplexes are reported. During the simulation the hexamers retained B‐type DNA structures that differed from typical A‐ or B‐DNA forms. The overall helical structures for the two duplexes are vary similar. The presence of G · T mispairs did not alter the overall helical structure of the oligonucleotide duplex. Large propeller twist and buckle angles were obtained for both duplexes. The purine/pyrimidine crossover step showed a large decrease in propeller twist in the normal duplex but not in the mismatch duplex. Upon the formation of wobble mispairs in the mismatched duplex, the guanines moved into the minor groove and the thymines moved into the major groove. This helped prevent purine/purine clash and created a deformation in the relative orientation of the glycosidic bonds. It also exposed the free O4 of the thymines in the major groove and N2 of the guanines in the minor groove to interactions with solvent and counterions. These factors seemed to contribute to the apparently higher rigidity of the mismatched duplex during the simulation.
Molecular dynamics simulations of small peptides: Dependence on dielectric model and pH
Authors: Daggett Valerie , Kollman Peter A. , Kuntz Irwin D.
Published: December 2004

There has been much interest recently in the structure of small peptides in solution. A recent study by Bradley and co‐workers [(1989) in Techniques of Protein Chemistry, Hugli, T. E., Ed., Academic Press, Orlando, FL, pp. 531–546; (1990) Journal of Molecular Biology, 215, pp. 607–622] describes a 17‐residue peptide that is stable as a monomeric helix in aqueous solution at low pH, as determined by two‐dimensional nmr and CD spectroscopy. They also have determined the helix content of the peptide as a function of pH using CD. We performed molecular dynamics simulations, with an empirical force field, of this peptide at low pH, with three different dielectric models: a linear distance‐dependent dielectric function (ε = R); a modified form [J. Ramstein and R. Lavery (1988) Proceedings of the National Academy of Science, USA, Vol. 85, pp. 7231–7235] of the sigmoidal distance‐dependent dielectric function of Hingerty and co‐workers [(1985) Biopolymers, Vol. 24, pp. 427–439]; and ε = 1 with the peptide immersed in a bath of water molecules. We found that simulations with the sigmoidal dielectric function and the model with explicit water molecules resulted in average distances for particular interactions that were consistent with the experimental nmr results, with the sigmoidal function best representing the data. However, these models exhibited very different helix‐stabilizing interactions. We also performed simulations using the sigmoidal function at moderate and high pH to compare to experimental determinations of the pH dependence of helix content. Helix content did not decrease with increases in pH, as shown experimentally. We did, however, observe changes in a specific side chain–helix dipole interaction that was implicated in determining the pH‐dependent behavior of this peptide. Overall, the sigmoidal dielectric function was a reasonable alternative to adding explicit water molecules. In comparing 100 ps molecular dynamics simulations, the sigmoidal function was much less computer intensive and sampled more of conformational space than the treatment using explicit water molecules. Sampling is especially important for this system since the peptide has been shown experimentally to populate both helical and nonhelical conformations.
Proton and tritium nmr relaxation studies of peptide inhibitor binding to bacterial collagenase: Conformation and dynamics
Authors: Dive Vincent , Lai Adolfo , Valensin Gianni , Saba Giuseppe , Yiotakis Athanasios , Toma Flavio
Published: December 2004

The interaction of succinyl‐Pro‐Ala, a competitive inhibitor of Achromobacter iophagus collagenase, with the enzyme was studied by longitudinal proton and tritium relaxation. Specific deuterium and tritium labeling of the succinyl part at vicinal positions allowed the measurement of the cross‐relaxation rates of individual proton or tritium spin pairs in the inhibitor–enzyme complex as well as in the free inhibitor. Overall correlation times, internuclear distances, and qualitative information on the internal mobility in Suc1 (as provided by the generalized order parameter S2) could be deduced by the comparison of proton and tritium cross‐relaxations of spin pairs at complementary positions in the CH2CH2 moiety as analyzed in terms of the model‐free approach by Lipari and Szabo.

The conformational and motional parameters of the inhibitor in the free and enzyme‐bound state were directly compared by this method. The measurement of proton cross‐relaxation in the Ala residue provided additional information on the inhibitor binding. The determination of the order parameter in different parts of the inhibitor molecule in the bound state indicates that the succinyl and alanyl residues are primarily involved in the interaction with the enzyme active site. The succinyl moiety, characterized in solution by the conformational equilibrium among the three staggered rotamers–i.e., trans: 50%; g+: 20%; g−; 30%–adopted in the bound state the unique trans conformation.
The Electrostatically Driven Monte Carlo method: Application to conformational analysis of decaglycine
Authors: Ripoll Daniel R. , Vásquez Max J. , Scheraga Harold A.
Published: December 2004

The Electrostatically Driven Monte Carlo (EDMC)method was applied in a study of a decamer of glycine whose conformational behavior is described by the Empirical Conformational Energy Program for Peptides (ECEPP/2) potential energy model. When free neutral end groups were used, it was found that conformations that were not α‐helical had significantly lower potential energies than fully α‐helical ones. However, when the N‐ and C‐termini were blocked by acetyl and methyl amide groups, respectively, the number of unsatisfied hydrogen‐bond donors and acceptors at the helix termini was diminished from 8 to 6; in this case, the possibility of forming two additional α‐helical hydrogen bonds was an important enough factor in making the α‐helical conformation the one with the lowest energy. The EDMC method was used as a global energy optimizer since it does not often become trapped in high‐energy local minima.
A method for estimating the nearest neighbor base‐pair content of RNAs using CD and absorption spectroscopy
Authors: Johnson Kenneth H. , Gray Donald M.
Published: December 2004

CD and absorption spectra are sensitive to the secondary structure of RNAs. By fitting the spectra contained in our basis set to the CD and absorption spectra of an RNA of known sequence, we could determine the fractions of base pairs, the fractions of each of the nearest neighbor base pairs, and the fractions of the single‐stranded nucleotides in that RNA. The basis set included 58 CD and 58 absorption spectra. The fitting was done with a guided selection routine. The estimated error was about 0.05 for predicting the fractions of the nearest neighbor base pairs, 0.06 for predicting the fractions of A · U, G · C, and G · U base pairs, and 0.04 for predicting the fractions of the single‐stranded nucleotides.
α‐Helical small molecular size analogues of neuropeptide Y: Structure‐activity relationships
Authors: Jung Günther , Beck‐Sickinger Annette G. , Dürr Hansjörg , Gaida Wolfram , Schnorrenberg Gerd
Published: December 2004

C‐terminal analogues of neuropeptide Y (NPY) of small molecular size have been synthesized. The influence of chain length, single or multiple amino acid substitution, and segment substitutions on receptor binding, pre‐ and postsynaptic biological activity, and conformational properties have been investigated. Receptor binding and in vivo assays revealed biological activity for NPY Ac‐25‐36 that increased with increasing α‐helicity. In attempts to stabilize the α‐helical content, three independent types of modified NPY Ac‐25‐36 analogues were synthesized. Strong agonistic activities could be detected in a series of discontinuous analogues, which are constructs of N‐terminal parts linked via different spacer molecules to C‐terminal segments. One of the most active molecules was NPY 1‐4‐Aca‐25‐36 (Aca, ε‐aminocaproic acid). For the first time conformational properties of a series of small NPY analogues have been investigated by CD, and correlated with biological activity and receptor binding. A C‐terminal dodecapeptide segment of NPY with an amount of 50% substitution to the native C‐terminal sequence of NPY was found to exhibit significant receptor binding.
Fluorescence, CD, and nmr studies on spantide, a bombesin and substance P antagonist
Authors: Di Bello Carlo , Scatturin Angelo , D'Auria Gabriella , Gargiulo Mario , Paolillo Livio , Trivellone Enrico , De Castiglione Roberto
Published: December 2004

The conformational properties of spantide [(D‐Arg1, D‐Trp7,9, Leu11)substance P] have been studied by fluorescence, CD, and nmr techniques. The fluorescence, CD, and nmr parameters in different solvents and in a micellar environment (SDS) are compared with the data collected for bombesin. A preliminary investigation on [D‐Pro2] spantide is also reported.
Ion binding of cyclolinopeptide A: An nmr and CD conformational study
Authors: Tancredi Teodorico , Benedetti Ettore , Grimaldi Maria , Pedone Carlo , Rossi Filomena , Saviano Michele , Temussi Piero Andrea , Zanotti Giancarlo
Published: December 2004

CD and nmr techniques have been used to study, in acetonitrile solution, the ion‐complexing capability of Cyclolinopeptide A (CLA), a cyclic nonapeptide of sequence

endowed with remarkable cytoprotective ability in vitro, and the conformation of the Ba2+/ CLA complex.

At room temperature, CLA in acetonitrile shows a proton nmr spectrum characteristic of the coexistence of many different conformers in intermediate exchange. The backbone contains a cis Pro‐Pro bond, with all other peptide bonds in the transconformation.

CLA binds Ba2+ more tightly than the other cations studied, namely K+, Na+, Mg2+, and Ca2+; CD data are indicative of the presence of both 1 : 2 (sandwich) and 1 : 1 (equimolar) type complexes, depending on the Ba2+ ion concentration, whereas nmr data are consistent with an equimolar form.

The relevant conformational features of the equimolar Ba2+/CLA complex are that the backbone contains all transpeptide bonds, a type I 6 → 3 β‐turn and a 3 → 1 γ‐turn (or a distorted 3 → 9 β‐turn). The global shape of the complexed peptide can be described as a bowl, with the concave (polar) side hosting Ba2+ and the convex side predominantly apolar.
Internal dynamics of linear and superhelical DNA as studied by photon correlation spectroscopy
Authors: Seils J. , Dorfmüller TH.
Published: December 2004

Photon correlation spectroscopy has been used to study the translational (D0)and internal dynamics of monodisperse linear and polydisperse superhelical plasmid‐DNAs. Scattering vector dependent correlation functions were measured and analyzed with the inverse Laplace transform CONTIN written by S. Provencher. For scattering vectors (q) lower than 1.3 × 105cm−1, D0 can be separated from internal dynamics. Using the measured D0value, the q dependence of internal modes was determined. Extrapolation of the internal relaxation times for q → 0 yields the longest internal relaxation time t1. This time can be connected to the persistence length in terms of the Berg‐Soda model, which describes the molecule as a semiflexible circular polymer with hydrodynamic interactions. The calculated length of 76 nm for DNA, I = 0.15 mol/L, is a little higher than the one obtained from static light scattering data without excluded volume corrections. A comparison of experimental and simulated correlation functions for the Berg‐Soda model shows that the model gives a fairly good description of the dynamics of the linear molecule, whereas large discrepancies between model and experimental functions are observed for the superhelical DNA. Small differences between model and experimental functions are mainly attributed to the neglect of the torsional modes that may be coupled to bending and flexing modes. For the superhelical DNA the agreement is improved if the calculation is carried out with a linear molecule, with shorter contour length and increased diameter. Both quantities can be derived from the known superhelix tilt angle.
Investigation of zinc‐binding affinities of moloney murine leukemia virus nucleocapsid protein and its related zinc finger and modified peptides
Authors: Mely Yves , Cornllle Fabrice , Fournié‐Zaluski Marie‐Claude , Darlix Jean‐Luc , Roques Bernard F. , Gérard Dominique
Published: December 2004

Nucleocapsid proteins of retroviruses are small basic, nucleic acid‐binding proteins with either one or two “Cys‐His” boxes, which have been shown to be involved in genomic RNA dimerization, encapsidation, and replication primer tRNA annealing to the initiation site for reverse transcription. The nucleocapsid (NC) protein of Moloney murine leukemia virus (MoMuLV NCp10) is made up of 56 residues with one Cys‐His motif. The Zn2+‐binding affinities and induced conformational changes of NCp10 were investigated by following the fluorescence of Trp 35 located in the Cys‐His domain. At pH 7.5, NCp10 was shown to bind Zn2+ at a 1 : 1 ratio with a very high apparent binding constant of 1.2 (±0.3) · 1013M−1. A similar apparent binding constant was obtained for a 19‐residue peptide encompassing the Cys‐His box, designated the “zinc finger motif,” indicating that it contains most if not all the information to bind Zn2+ tightly. Changing Trp 35 to Phe in the peptide did not affect the Zn2+ affinity, indicating that Trp 35 is not crucial for Zn2+ binding. On the contrary‐, replacing Cys 29 by Ser, the chemical modification or oxidation of the three Cys sharply reduced Zn2+ affinity, confirming the essential role of Cys in Zn2+ binding. In addition, fluorescence and energy transfer data suggested that Zn2+ binding modifies the Trp 35 environment but not its solvent exposure, and increases the average distance between Tyr 28 and Trp 35 by about 2 Å. These data suggest that Zn2+ binding to retroviral NC protein is biologically relevant.
Effects of limited input distance constraints upon the distance geometry algorithm
Authors: Oshiro C. M. , Thomason John , Kuntz I. D.
Published: December 2004

In this paper we examine the distance geometry (DG) algorithm in the form used to determine the structure of proteins. We focus on three aspects of the algorithm: bound smoothing with the triangle inequality, the random selection of distances within the bounds, and the number of distances needed to specify a structure. Computational experiments are performed using simulated and real data for basic pancreatic trypsin inhibitor (BPTI) from nmr and crystallographic measurements. We find that the upper bounds determined by bound smoothing to be a linear function of the true crystal distance. A simple model that describes the results obtained with randomly selected trial distances is proposed. Using this representation of the trial distances, we show that BPTI DG structures are more compact than the true crystal structure. We also show that the DG‐generated structures no longer resemble test structures when the number of these interresidue distance constraints is less than the number of degrees of freedom of the protein backbone. While the actual model will be sensitive the way distances are chosen, our conclusions are likely to apply to other versions of the DG algorithm.
Intercalation binding of 6‐substituted naphthothiopheneamides to DNA: Enthalpy and entropy components
Authors: Hopkins Harry P. , Ming Yang , Wilson W. David , Boykin David W.
Published: December 2004

N‐(3‐dimethylaminopropyl) naphtho [2, 1‐b] thiophene‐4‐carboxamide and the 6‐substituted methoxy, methyl, fluoro, chloro, bromo, trifluoromethyl, and cyano derivatives have been shown to bind to DNA via intercalation with binding constants in the 35–900 × 103 range at 25°C, pH 7, and [Na+] = 0.019M . Both electron‐donating and ‐withdrawing substituents enhance intercalation binding, but the binding affinity is most enhanced by the cyano substituent. Calorimetric titrations for calf thymus DNA differ dramatically from those reported for ethidium [Hopkins et al. (1990) Biopolymers Vol. 29, pp. 449–459]. Apparent enthalpy parameters (ΔHB) for intercalation are constant only at low coverage of sites and become much more positive as saturation is approached. In the plateau region, ΔHB values for the parent and the cyano‐, fluoro‐, chloro‐, and bromo‐substituted compounds are nearly the same (∼ −5.9 kcal/mol). For the methyl‐ (−6.8 kcal/mol) and methoxy‐ (−7.5 kcal/mol) substituted compounds, the ΔHB values are more exothermic than that for the un‐substituted compound, whereas ΔHB for the trifluoromethyl compound is approximately 1 kcal/mol less exothermic. The corresponding ΔSB values, corrected for mixing effects, are in the 7–15‐cal/deg/mol range and are approximately linearly related to ΔHB if the cyano derivative is excluded.
Electrostatic interactions in protein solution—a comparison between poisson–boltzmann and Monte Carlo calculations
Authors: Fushiki M. , Svensson B. , Jönsson Bo , Woodward C. E.
Published: December 2004

The accuracy of the Poisson–Boltzmann (PB) approximation and its linearized version is investigated by comparison to results obtained from Monte Carlo simulations. The dependence of the calcium binding constant of the protein calbindin as a function of salt concentration and mutation is used as a test case. The protein is modeled as a collection of charged and neutral spheres immersed in the electrolyte solution. The PB equation is solved using a finite difference technique on a grid in a spherical polar coordinate system, which is the preferred choice for a globular protein like calbindin. Both MC and PB give quantitative agreement with experimental results. The linearized PB equation is almost as accurate, but it becomes less reliable in systems with divalent ions. However, the linearized PB equation fails to describe the concentration profiles for cations and anions outside the protein even in a 1 : 1 salt solution.
Conformational analysis of a IgG1 hinge peptide derivative in solution determined by NMR spectroscopy and refined by restrained molecular dynamics simulations
Authors: Kessler Horst , Mronga Siggi , Müller Gerhard , Moroder Luis , Huber Robert
Published: December 2004

The hinge region links the antigen binding Fab part to the constant Fc domain in immunoglobulins. For the hinge peptide derivative [AcThr(OtBu)‐Cys‐Pro‐Pro‐Cys‐Pro‐Ala‐ProNH2]2 the assignment of the 1H and 13C resonances was achieved by two‐dimensional nmr techniques: total correlation spectroscopy (TOCSY), nuclear Ovethauser enhancement spectroscopy (NOESY), rotating frame nuclear Overhauser enhancement spectroscopy (ROESY), heteronuclear multiple quantum coherence (HMQC) transfer, and a HSQC (modified Overbodenhausen experiment) with high resolution in F1, which was several times folded in F1 but still phase correctable. Conformational relevant parameters (78 nuclear Overhauser effect distance restraints, 3JHH for prochiral assignments, temperature gradients) were determined by nmr and served as input data for molecular dynamics (MD) structure refinement. A simulated model compound corresponding to the [Cys‐Pro‐Pro‐Cys]2 core elongated by the peptide chains in the Fab and Fc direction served as a starting structure for the final MD run. The conformation calculated in in vacuo does not agree with the C2 symmetry required from nmr data, but the structure obtained by a water simulation fulfills the requirement. Here the core of the hinge peptide derivative adopts a polyproline II double helix as in the x‐ray structure of IgG1. Hence, segments responsible for the internal flexibility are located outside the core as confirmed by the flexibility of the solvent exposed C termini.
Comparison of the bis‐intercalating complexes formed between either ditercalinium or a flexible analogue and d(CpGpCpG)2 or d(TpTpCpGpCpGpApA)2 minihelices: 1H‐ and 31P‐nmr analyses
Authors: Pothier Joël , Delepierre Muriel , Barsi Marie‐Christine , Garbay‐Jaureguiberry Christiane , Igolen Jean , Le Bret Marc , Roques Bernard P.
Published: December 2004

The 400‐MHz 1H‐ and 162‐MHz 31F‐nmr have been used to study complexes constituted by (a) the d(TpTpCpGpCpGpApA)2 or the d(CpGpCpG)2 self‐complementary oligonucleotides and (b) two bifunctional 7H‐pyrido [4, 3‐c] carbazole dimer drugs, the antitumoral ditercalinium (NSC 366241), a dimer with a rigid bis‐piperidine linking chain and its pharmacologically inactive analogue, a dimer with a flexible spermine‐like linking chain. Nearly all proton and phosphorus signals have been assigned by two‐dimensional (2D) nmr (correlated spectroscopy, homonuclear Hartmann‐Hahn, nuclear Overhauser enhancement spectroscopy, 2D 31P {1H} heteronuclear correlated spectroscopy and 31P‐31P chemical exchange experiments). Both drugs bis‐intercalate into the two CpG sites. The complexes show small differences in the position of the 7H‐pyrido[4, 3‐c]carbazole ring into the intercalation site and possibly in the ribose‐phosphate backbone deformation. However, the inactive analogue exhibits a longer residence lifetime in octanucleotide than the ditercalinium does. All these results are discussed in terms of differences in dimer activities.
Resolution of end‐to‐end diffusion coefficients and distance distributions of flexible molecules using fluorescent donor‐acceptor and donor‐quencher pairs
Authors: Lakowicz Joseph R. , Kuśba Józef , Szmacinski Henryk , Gryczynski Ignacy , Eis Peggy S. , Wiczk Wieslaw , Johnson Michael L.
Published: December 2004

We used time‐dependent fluorescence energy transfer, time–dependent collisional quenching, and global analysis of the data resulting from these through–space and contact interactions to recover the end–to–end distance distributions and diffusion coefficients of flexible fluorescent molecules. The fluorescence decays of covalently linked tryptamine–acceptor and tryptamine–quencher pairs were measured by the frequency–domain method. These data were fit using numerical solutions of the differential equation, which predicts the time–and distance–dependent population of the excited state donors in the presence of energy transfer or collisional quenching, followed by transformation to the frequency domain for nonlinear least–squares comparison with the experimental data. We found that the energy transfer data for the donor–acceptor pair alone were adequate to recover the starting distribution and the end–to–end diffusion coefficient; however, the resolution is dramatically improved by the use of both the through–space and contact interactions.
Kinetics of folding and unfolding of αα‐tropomyosin and of nonpolymerizable αα‐tropomyosin
Authors: Mo Jianming , Holtzer Marilyn Emerson , Holtzer Alfred
Published: December 2004

Stopped flow CD (SFCD) kinetic studies of self–assembly of coiled coils of rabbit αα–tropomyosin and of nonpolymerizable αα–tropomyosin (NPTm) are reported. The protein was denatured in 6M urea buffer, then renatured by 10‐fold dilution into benign saline buffer. Folding was monitored by SFCD in the backbone region (222 nm). Protein chains are shown to be totally unfolded (and separated in the reduced species) in the initial denaturing medium and fully folded as two‐chain coiled coils in the final benign medium. In all cases of folding in benign buffer of totally unfolded chains, two phases were found in the folding process: a fast phase (<0.04 s, the SFCD dead time), in which an intermediate state with about 70% of the equilibrium ellipticity forms; followed by a slower, observable phase that completes the folding. The slow phase is first order (k −1 = 1.6 s at 20°C), signifying that chain association for reduced samples occurs in the fast phase. In contrast, folding in benign buffer from an initial state with 70% of the equilibrium ellipticity is all fast, suggesting that the folding intermediate is not an equilibrium species. Cross‐linking at Cys‐190 increases the helix content of the fast‐formed intermediate state to about 85% of the equilibrium value, but leaves the rate constant of the slow phase unchanged. In NPTm, which does not form high aggregates at low ionic strength, the rate of the observable phase is almost independent of ionic strength in the range of ∼ 0.15‐0.6M, but is reduced one to two orders of magnitude by further reduction to 0.026M. In folding from totally unfolded chains, the rate is reduced less than one order of magnitude by changing the final state to about 50% folded. In contrast to folding, unfolding of αα‐tropomyosin from the native state is all fast.
Photodegradation of hyaluronic acid: EPR and size exclusion chromatography study
Authors: Lapc̆ík L'. , Chabrec̆ek P. , Stas̆ko A.
Published: December 2004

Photochemically induced radical reactions involving the lateral sequences and the end macromolecular chain groups of hyaluronic acid in aqueous solutions at 293K were studied by EPR spin trapping technique with DMPO (5,5‐dimethylpyrroline‐1‐oxide). In the first 1‐10 minutes of irradiation EPR indicates spin adducts of two carbon centered radicals with the splitting constants of aN = 1.60 mT, aH = 2.51 mT and aN = 1.56 mT, aH = 2.28 mT. After longer irradiation time (over 10 minutes) dominate two further DMPO adducts of radicals centered on hetero–atoms with splitting constants of aN = 1.44 mT, an = 1.60 mT and of aN = 1‐49 mT, aH = 1.49 mT. Simultaneously, molecular weight followed by SEC decreases, suggesting that UV irradiation leads to the breaking of interglycosidic bonds of hyaluronic acid main macromolecular chain.
Development of an extended simulated annealing method: Application to the modeling of complementary determining regions of immunoglobulins
Authors: Higo Junichi , Collura Vincent , Garnier Jean
Published: December 2004

An extended simulated annealing process (ESAP) has been developed in order to obtain an ensemble of conformations of a peptide segment from a protein fluctuating at a given temperature. The annealing process was performed with a fast Monte Carlo method using the scaled collective variables developed by Noguti and Gō. The system was divided into two parts: one consists of one or more peptide segments and is flexible around the main‐chain and side‐chain torsional angles; the other represents the rest of the molecule and was maintained fixed at the atomic positions determined by x‐ray experiments. The target function included the nonbonding atomic interactions and a distance function to anchor the N and C terminal ends of each segment to the fixed part. Three systems of complementary determining regions (CDR) of antibodies were tested and compared to x‐ray data: L2 loop (7 residues) of the light chain of α‐type Bence‐Jones protein, H1 and the H2 loops (14 residues) of McPC603, and H1 and H2 loops (12 residues) of HyHEL‐5. Each state of CDR conformations was characterized at room temperature by the average of their coordinates (average conformation) and the internal energy. With a limited number of annealing processes (10), starting from the extended conformation, we have obtained states with conformations close to the observed x‐ray structures, from 1.1 to 1.7 Å root mean square deviation (rmsd) of main‐chain atoms depending on the system. These states were identical or within 0.25 Å rmsd of those of lowest internal energy. For unknown CDR structures the criteria of lowest internal energies from ESAP can be used to predict hypervariable loop structures in antibodies with an accuracy comparable to other methods.