bluuebunny/crossref_metadata_2025 huggingface dataset:
Dielectirc relaxation of DNA in aqueous solutions
Authors: Sakamoto Masanori , Kanda Hirohsi , Hayakawa Reinosuke , Wada Yasaku
Published: May 2005

Using a four‐electrode cell and a new electronic system for direct detection of the frequency differences specturm of solution impedance, the complex dielectric constant of calf thymus DNA (Mr = 4 × 106) in aqueous NaCl at 10°C is measured at frequencies ranging from 0.2 Hz to 30 kHz. The DNA concentrations are Cp = 0.01% and 0.05%, and the NaCl concentrations are varied from Cs = 10−4 M to 10−3 M. A single relaxation regions is found in this frequency range, the relaxation frequency being 10 Hz at Cp = 0.01% and Cs = 10−3 M. At Cp = 0.05% it is evidenced that the DNA chains have appreciable intermolecular interactions. The dielectric relaxaton time τd at Cp = 0.01% agrees well with the rotational relaxation time estimated from the reduced visocisty on the assumption that the DNA is not representable as a rigid rod but a coiled chain. It is concluded that the dielectric relaxiatioinis ascribed to the rotation of the molecule. Observed values of dielectric increment and other experimental findings are reasonably explained by assuming that the dipole moment of DNA results from the slow counterion fluctuation which has a longer relaxation time than τd.
Electric field‐induced conformational change of poly(L‐lysine) studied by transient electric birefringence
Authors: Kikuchi Kazuo , Yoshioka Koshiro
Published: May 2005

Transient electric birefringence measurements on poly(L‐lysine hydrobromide) in methanol–water mixtures have been carried out at various solvent compositions in the vicinity of the helix–coil transition region (from 87 to 98 vol % methanol). Anomalous birefringence transients were observed between 90 and 95 vol % methanol above a threshold field strength. A distinct difference between the responses to weak and strong electric fields was noticed over a narrow range of the solvent composition. The effects of polymer concentration and temperature on the field‐strength dependence of the birefringence were studied at a solvent composition of 90 vol % methanol where the anomalous transients appeared most clearly. The double logarithmic plots of the steady‐state specific birefringence versus the square of field strength for different concentrations and temperatures could be superimposed by shifting them horizontally along the abscissa. The threshold field strength which was determined from the shift factor decreased with decreasing concentration. The results provide further evidence that strong electric fields can cause a helix–coil transition in this system under favorable conditions.
Induced circular dichroism of acridine orange bound to double‐stranded RNA and transfer RNA
Authors: Zama Mitsuo , Ichimura Sachiko
Published: May 2005

The induced circular dichroism (CD) in the visible region of acridine orange bound to the double‐stranded RNA from cytoplasmic polyhedrosis virus and to yeast tRNA has been measured as a function of RNA phosphate‐to‐dye ratio (P/D), under the conditions of 0.01 M Na+ at pH 7.0. The shape of the CD spectrum of acridine orange bound to the double‐stranded RNA was quite different from the spectrum of the dye bound to DNA. The CD spectral features of acridine orange bound to the double‐stranded regions in tRNA closely resembled those of the double‐stranded RNA‐dye complex, suggesting that the dyes bind similarly to the two RNA's. It was further found that the CD spectrum of the tRNA‐dye complex at sufficiently high P/D ratios, which is assignable to monomeric, intercalated dye to the base‐paired parts in tRNA, is also distinct from the corresponding spectrum of the DNA‐dye complex.
The spatial distribution of moving macromolecules undergoing isomerization
Authors: Mitchell R. M.
Published: May 2005

In this paper we use probabilistic arguments to derive and discuss the spatial distribution of molecules which are undergoing electrophoresis or centrifugation while at the same time they are switching back and forth between two configurational states. An exact solution is obtained for arbitrary values of the diffusion coefficients of the two isomerizing states. The traditional analytic method of solving this problem, namely, by use of “master equations,” is not completely satisfactory because one can find only the Fourier transform of the solution rather than the solution itself. Our treatment yields the answer one would get if it were feasible to perform the inversion of the Fourier transform.
An easy method for retrieving atomic coordinates from CPK space‐filling molecular models
Authors: De Haën Christoph , Swanson Eric , Teller David C.
Published: May 2005

Connectors for CPK space‐filling molecular models have been supplemented with pointed steel wires (ϕ = 1.59 mm) of appropriate lengths and positions to represent atomic bonds. For illustration purposes, the polypeptide backbone of glucagon was equipped with such wire‐core connectors and given a possible conformation. The molecular model was packed into a cubic box and, using a clinical instrument, X‐ray films were taken from three orthogonal directions. Atomic coordinates on the films were then corrected to real‐space coordinates.

This method for retrieving atomic coordinates from space‐filling models of modium‐sized polypeptides is easy and can be done wih equipement available everywhere. Coordinates obtained in this way should be useful as starting points for energy minimization, for the calculation of frictional coefficients, and in attempts to solve crystal structures by X‐ray diffraction, using molecular replacement methodology.
Kinetics of proflavin binding to bacteriophages T2L and T4D
Authors: McCall Philip J. , Bloomfield Victor A.
Published: May 2005

We have measured the kinetics of proflavin binding to T‐even bacteriophages—the 700 S and 1000 S forms of T2L, T4D, and T4D os41—by difference spectroscopy at 430 nm. Measurements were carried out from 22° to 37°C. Binding is very slow to encapsulated DNA compared to free DNA, requiring hours to reach equilibrium. The kinetic data are compatible with the two‐step mechanism

where P is proflavin, N is nucleotide, and PN1 and PN2 are complexes. Computer integration of the rate equations allows evaluation of the rate constants; previous equilibrium measurements gave thermodynamic parameters. For all phage studied, the bimolecular step is endothermic with high positive entropy; the second, unimolecular step is highly exothermic with small negative entropy change. Both forms of T2L bind proflavin with essentially the same rate, as do T4D and the osmotic shock resistant mutant T4D os41. This suggests that the encapsulated DNA is equally accessible to proflavin in both forms of each phage. However, T4D binds dye appreciably faster than T2L, indicating that capsid permeability or DNA environment (glucosylation or packing) is different in the two species.
Vibrational analysis of peptides, polypeptides, and proteins. I. Polyglycine I
Authors: Moore W. H. , Krimm S.
Published: May 2005

A force field has been refined for the antiparallel chain‐rippled sheet structure of polyglycine I. Transition dipole coupling and hydrogen bonding are explicitly taken into account. Amide I and amide II mode splittings are well accounted for, the latter also providing a quantitative explanation of the amide A and amide B mode frequencies and intensities. In addition to predicting other features of the vibrational spectrum of polyglycine I, this force field is completely transferable to other β polypeptides, even though these have the antiparallel chainpleated sheet structure.
The optical properties of tyrosine‐containing cyclic dipeptides
Authors: Snow Julian W. , Hooker Thomas M. , Schellman John A.
Published: May 2005

The chiroptical properties of the cyclic dipeptides cyclo (L‐alanyl‐L‐tyrosine) and cyclo(L‐tyrosyl‐L‐tyrosine) have been investigated as a function of molecular conformation. Theoretical optical calculations and conformational energy calculations have been carried out as a function of the side‐chain dihedral angles χ1 and χ2, and as a function of the angle of fold of the cyclic dipeptide backbone. The results of these theoretical calculations have been compared with experimental circular dichroism (CD) data. Theoretical predictions are in very good agreement with experiment for c(L‐Tyr‐L‐Tyr). Agreement was not quite as good for c(L‐Ala‐L‐Tyr), although the signs of all of the Cotton effects were apparently predicted correctly except for that associated with the lowest energy tyrosine absorption band.
Experimental and theoretical investigations on the folding modes of depsipeptide molecules
Authors: Boussard G. , Marraud M. , Neel J. , Maigret B. , Aubry A.
Published: May 2005

The coupling of N‐acyl‐α‐amino‐acids with α‐hydroxyacid‐methyl amides results in depsipeptide molecules containing two chiral centers and one ester function inserted between two amide functions. Their conformational features have been investigated by IR spectroscopy, proton magnetic resonance, X‐ray diffraction, and theoretical P.C.I.L.O. calculations.

It is shown that most of these molecules are folded by an intramolecular 4 → 1 hydrogen bonding. Two folded conformations, similar to the well known β turn in peptides, are described, the stability of which depends on the configurational sequence in the investigated molecule.

LL and LD species are folded in two different ways whereas LG sequences containing an achiral hydroxy‐acid residue accommodate both of them. The presence of a N‐terminal achiral amino acid noticeably decreases the folding ratio.

The above conclusions are then compared with the conformational features of homologous tripeptide molecules.
The organization and repair of DNA in the mammalian chromosome. I. Calibration procedures and errors in the determination of the molecular weight of a native DNA
Authors: Lange Christopher S. , Liberman Daniel F. , Clark Roger W. , Ferguson Pamm
Published: May 2005

The semiautomated sucrose gradient system of Lange and Liberman (1974) (Anal. Biochem. 59, 129–145) has been used to determine an accurate value of the exponent k of the Burgi and Hershey equation (1963) (Biophys. J. 3, 309–321) under conditions of 1M salt. A complete analysis of the variance of k was done for data from both systems. The results lead to the conclusion that the value k = 0.401 (±0.012 for reproducibility; ±0.019 for total error) obtained here is considerably more accurate than that of Burgi and Hershey (even when corrected for DNA concentration effects and the improved size estimates of T4 DNA size). The use of the Burgi and Hershey relationship and a T4 DNA metric to determine molecular weights of mammalian DNA requires the utmost precision in the exponent if useful estimates are to be obtained. This also has implications for double‐strand breakage and repair estimates.
A study of paramyosin aggregation using transient electric birefringence techniques
Authors: Krause Sonja , DeLaney Donald E.
Published: May 2005

The aggregation of several forms of the molluscan muscle protein paramyosin at low concentration and at low ionic strength was studied in the pH range 6–10 using transient electric birefringence techniques. In the lower part of this pH range, the aggregates exhibit negative birefringence, but this changes to positive birefringence when the pH increases. Analysis of the field‐free birefringence decay transients of the paramyosin solutions showed that all aggregates coexisted with paramyosin monomer and allowed a determination of the rotational diffusion constant of the aggregates present at each pH. The size and shape of these aggregates were estimated from their rotational diffusion constants and were compared with the known characteristics of larger aggregates such as paracrystals. The positively birefringent aggregates appear to be staggered dimers at certain values of pH; at other pH values, these aggregates appear to be higher aggregates, probably formed by lateral aggregation of monomer or dimer onto one of these staggered dimers. The staggered dimers are formed by overlap of 200–600 Å along each cylindrical paramyosin molecule, in agreement with the 530‐Å overlap distance found in paramyosin paracrystals by Cohen et al. (1971) J. Mol. Biol. 56, 223–237. Some speculations are presented about the nature of the negatively birefringent species.
Intramolecular and overall motion of proline: The influence of viscosity on carbon‐13 spin‐lattice relaxation times
Authors: Deslauriers Roxanne , Smith Ian C. P.
Published: May 2005

Nuclear magnetic resonance of 13C is used to probe the overall and internal motions of proline. Spin‐lattice relaxation times (T1) are reported for proline monomer dissolved in water/glycerol mixtures. Rates of overall molecular motion and internal motion depend on solvent composition but to different degrees. The effective correlation times (τeff) of the various proton‐bearing carbon atoms in proline vary linearly as a function of solvent composition (%v/v) rather than of solution viscosity. The effective correlation time for molecular motion (τeff) is separated into contributions from overall molecular motion (τmol) and internal motion (τint). The γ‐carbon of proline shows the smallest dependence of τint on solvent composition. The data indicate a high degree of intramolecular motion for the γ‐carbon of proline. Inclusion of anisotropic molecular reorientation in the data analysis was found not to affect the above conclusions. The observed values of τeff indicate that the rotational diffusion model of molecular reorientations should apply to proline. The values of τeff calculated for proline using the Stokes‐Einstein relation are larger than those observed; the discrepancy is discussed in terms of solvent‐solute interactions.
1H nuclear magnetic resonance studies of N‐acetyl‐L‐proline N‐methylamide. Molecular conformations, hydrogen bondings, and thermodynamic quantities in various solvents
Authors: Higashijima Tsutomu , Tasumi Mitsuo , Miyazawa Tatsuo
Published: May 2005

Concentration and temperature dependences of the 1H nmr spectra of N‐acetyl‐L‐proline N‐methylamide were observed in various solvents [CCl4, CDCl3, (CD3)2CO, (CD3)2SO, H2O, and D2O]. The fraction of the cis isomer (with respect to the bond between the acetyl carbonyl carbon and prolyl nitrogen atoms) depends greatly on the solvent used; the fraction of the cis isomer is higher in polar solvents than in nonpolar solvents. It depends also on concentration and temperature in nonpolar solvents but not in polar solvents. In nonpolar solvents the trans isomer mostly exists in the γ‐turn structure with an intramolecular hydrogen bond and the cis isomer tends to form molecular aggregates by intermolecular hydrogen bonds. In polar solvents both the cis and trans isomers exist in monomeric forms which interact with solvent molecules. The pH dependences of the N‐methyl proton resonances indicate that the γ‐turn structure of the trans isomer is present also in aqueous solution, though its population is difficult to determine. Apparent enthalpy and entropy changes for the conversion of the trans isomer to cis isomer are evaluated for various solvents. The results are discussed in terms of the intra‐ and intermolecular hydrogen bondings.
Cross linking of proteins in vitro by peroxidase
Authors: Stahmann M. A. , Spencer A. K. , Honold G. R.
Published: May 2005

The enzyme peroxidase, a substrate (hydrogen donor), and hydrogen peroxide aggregated and polymerized soluble proteins included in the reaction mixture. Gel filtration and acrylamide disk gel electrophoresis revealed newly formed dimers, trimers, and higher protein polymers. Some of the protein polymers withstood the denaturing conditions of dodecyl sulfate disk gel electrophoresis; thus the formation of some covalent cross links was indicated. It is suggested that peroxidase catalyzes the oxidation of hydrogen donors to form free radicals or quinones, which subsequently interact with, cross link, and alter the soluble proteins.
Quantum yields and fluorescence lifetimes of acridine derivatives interacting with DNA
Authors: Duortail Guy , Mauss Yves , Chambron Jacques
Published: May 2005

Interactions of several acridine dyes with DNA from different species were studied by measuring fluorescence lifetimes in the 2–30‐nsec range, using the single‐photon counting technique, and by measuring fluorescence quantum yields in the steady state. The results confirm the existence of two principal site classes, one in which the dye fluorescence is quenched by interaction with guanine and another in which fluorescence results from the hydrophobic environment of the A·T base pairs. The emitting sites are found, in some cases, to exhibit fluorescent decay curves which can be resolved into two exponential components corresponding to a short and to a long lifetime. The deviation from one exponential component is particularly clear with rivanol, 9‐aminoacridine, and quinacrine, with which one component is two or three times longer than the other. The relative proportion of these two components depends only slightly on the DNA base composition and does not depend on the nature of the acridine derivatives. We postulate that this lifetime heterogeneity corresponds to the two discrete steps in the complex formation elucidated by kinetic studies: the first step corresponds to a semi‐intercalated, or “external,” dye with a short fluorescence lifetime and the second step corresponds to a totally intercalated dye with a long lifetime. In this model, we assumed that a transient opening of the site near a semi‐intercalated dye induces solvent diffusion which in turn is responsible for its short‐lived fluorescence.
Helical segment analysis of α‐helical regions in proteins
Authors: Rajan S. S. , Srinivasan R.
Published: May 2005

The methods suggested earlier for the analysis and representation of protein structural data are now extended to the helical regions in finer details. These enable better handling of characterization of bends and distortions, for which statistical parameters are also developed. Using latest myoglobin data, best experimental parameters for the α‐helix are deduced to be rN = 1.55 (0.13) Å, r = 2.28 (0.12) Å, rC′ = 1.70 (0.10) Å, r0 = 2.02 (0.12) Å, ϕ = 100.5 (2.3)°, and t = 1.495 (0.055) Å.
Helix‐coiled transition in heteropolymers. I. Ground‐state energy
Authors: Vilenkin A.
Published: May 2005

A new technique is presented for treating the ground state of an heteropolymer with a random sequence of components. An exact system of equations is found for determining the ground state energy E which is equal to the polymer free energy f in the lowest‐order approximation in T/V (V/2 is the large “surface” energy arising at the boundaries between coiled and “helical” sections: V ≫ T, Uk; U1 and –U2 are the free energies of the components counted from the corresponding coiled state energies). These equations are essentially simplified at certain fixed values of the ratio U1/U2. For integer values of U2/U1 and U1/U2 a solution is obtained with an accuracy exp(–V/Uk). The ground‐state energy as a function of U1 and U2 is shown to be highly irregular: its derivatives have jumps at an infinite number of points. These jumps provide a fine structure of the melting curves. A smoothed over the jumps function E′ is found by way of analytic continuation from the integer values of U1/U2 and U2/U1. The accuracy of the approximation f ≈ E is estimated and the correctional term of order T/V is determined.
Extrinsic cotton effect and helix‐coil transition in a DNA‐polycation complex
Authors: Bhat G. , Roth A. C. , Day R. A.
Published: May 2005

A strong, positive, extrinsic CD band ([θ]242.5 = ∼2 × 10−3 deg cm2/dmole) has been observed for a α‐bromo‐poly[methylene‐1,4‐phenylenecarbonyloxyethylene(dimethylamino) bromide] (I). The extrinsic Cotton effect is attributed to the ordered arrangement of the aromatic chromophores along the DNA helix.

The extrinsic band had a linear dependence on the amount of polycation I added from r ≤ 0.3 to r = ∼0.5, but decreased thereafter. Addition of the polycation decreased the positive CD band of DNA at 275 nm. The transformation of B → C form in the presence of salts or other polycations caused similar changes. The decrease in [θ]275 was reversed at higher concentrations of the polycation (r > 0.4).

Thermal denaturation studies indicated both stabilization of the helix conformation (Δtm = 21°C) and a high degree of cooperativity in the melting of DNA‐polycation complex as compared to native calf thymus DNA. Using the linear relationship between r (polycation residue/DNA phosphate) and F (fraction of bound base pairs), a value of 0.6 was derived for β (number of monomer residues of polycation/nucleotide).

Both electrostatic and hydrophobic effects probably influence the stability of the DNA‐polycation complex, since the strength of the 242.5 nm CD band is a function of both salt and urea concentrations.
Water in keratin: Study of the depolarization thermal current peak II
Authors: Leveque J. L. , Garson J. C. , Boudouris G.
Published: May 2005

The depolarization thermal current method is applied to the study of the polarization phenomena in the keratin‐water system. Three depolarization thermal current peaks have been evidenced in hair keratin. This paper deals with the detailed study of peak II. For the first time, the mechanism responsible for this peak is ascribed to molecular reorientation. The effect of water upon the characteristics of peak II is also described, and an activation energy of 8.4 kcal/mol is computed. These results, in connection with other studies, lead us to interpret peak II as due to the reorientation in the bound or intermediate water molecules. The effect of copper confirms that the carboxyl groups are hydration sites.
Determination of the base‐pairing content of four specific tRNAs by infrared spectroscopy
Authors: Schernau U. , Ackermann Th.
Published: May 2005

Infrared spectroscopy was used for the determination of the base‐pairing content of four specific tRNAs in deuterium oxide solution. Infrared spectra were obtained in the 1750–1550 cm−1 region at various temperatures ranging from about 15 to 90°C. Melting curves were constructed by plotting the molar extinction coefficient at ν = 1657 cm−1 versus temperature. These transition curves enabled us to determine the ranges of temperature which correspond to the ordered (partially double‐stranded) or randomly coiled structure of the tRNA. For a set of wavenumbers the extinction coefficients at these temperatures were used for the calculation of the base‐pairing content. The procedure employed here is based on a method described earlier by Thomas [(1969) Biopolymers 7, 325–334]. For the conditions selected for this investigation (Mg2+‐free D2O‐buffer; 0.01M tris‐DCl, 0.015M NaCl, pD 7.5) the results of this determination agree within the limits of errors with the number of base pairs predicted by the cloverleaf model.
Hydrodynamics of macromolecular complexes. III. Bacterial viruses
Authors: De La Torre Jose Garcia , Bloomfield Victor A.
Published: May 2005

We have calculated the translational and rotational frictional coefficients of structures related to T2 and T4 bacteriophage, using the theoretical framework developed in the preceding two papers. The structures considered were models for tail‐fiberless phage, and for whole phage with fibers in the extended and retracted portions. We also computed and compared with the experiment the changes in translational frictional coefficient produced by successive addition of 1–6 fibers to the fiberless particle. Agreement with experimental results is markedly improved over previous theoretical efforts, especially with respect to the effect of tail‐fiber extension. Some significant discrepancies remain, however, in the comparison of fiber‐retracted and fiberless phage.
Rotational isomerism of N‐acetylamino acid methylamides having serine, tyrosine, histidine, and proline residues as revealed by Raman spectroscopy
Authors: Koyama Yasushi , Uchida Hisaaki , Oyama Shigeru , Iwaki Tetsuo , Harada Kimihiko
Published: May 2005

Rotational isomerism of N‐acetylamino acid methylamides having serine, tyrosine, histidine, and proline residues was investigated by means of Raman spectroscopy. The Raman spectra of each compound for solid states (crystalline or glassy) as well as for aqueous solution were compared with each other. Different kinds of rotational isomers were found to exist in the different solid states; three such solid states for N‐acetylserine methylamide, two for N‐acetyltyrosine methylamide, and one for N‐acetylhistidine methylamide were found in the present investigation. Additional rotational isomers were found to exist in aqueous solutions. The results indicated that the inherent relative stability of the rotational isomers differs little and is very dependent on intermolecular interaction, especially that due to hydrogen bonds. The possible molecular conformations are discussed on the basis of x‐ray results now available both for the present compounds and for proteins.
Quasielastic light scattering: Effect of ionic strength on the internal dynamics of DNA
Authors: Caloin Michel , Wilhelm Bernadette , Daune Michel
Published: May 2005

Quasielastic light scattering is used to study the effect of ionic strength on the dynamic behaviour of DNA. In a first approach the spectrum of scattered light is analyzed in terms of a single relaxation process. The large difference between the observed behaviour and that expected according to a pure diffusional process reflects the contribution associated with internal modes, which increases with decreasing ionic strength. Such behaviour is better analyzed in terms of a double relaxation process by using two relaxation times, the reciprocals of which are equal to DK2 and DK2 + τi−1 (K), respectively, where τi (K) is an average value describing the set of modes observed at a given K value. Relative intensity and relaxation times, which are the more accurate parameters, were used to interpret the results. The observed increase of the relative contribution of internal modes with decreasing ionic strength is actually a relative decrease of the diffusional contribution induced by a corresponding increase of the radius of gyration RG. On the other hand, the reciprocal τi−1 (K) of the relaxation time is a linear function of K2 in the analyzed KRG range and is insensitive to ionic strength between 10−2M and 1M. These results, when discussed according to Rouse's model, lead to define for each value of τi−1 (K) a corresponding mean‐squared equilibrium length 〈μ〉 which is found to be a linear function of K−2.
Correlation of sequence and tertiary structure in globular proteins
Authors: Crippen Gordon M.
Published: May 2005

The x‐ray crystal structures of 19 selected proteins are examined empirically for correlations between the amino acid sequence and long‐range, tertiary conformation. There is clear evidence for preferential associations of certain types of amino acids, particularly among the hydrophobic aliphatic, aromatic, and cysteine residues. However, the likelihoods of forming these residue‐pair contacts are all less than 12%, so packing and geometric requirements must often take precedent over energetic considerations. The prediction of long‐range contacts is not substantially improved by taking into account the sequentially previous residues. The analysis of atom–atom contacts shows a similar lack of predictive ability, but the results show that a good approximation to the interresidue energy function must include different types of interactions at two or three different sites on some amino acids. Backbone–backbone long‐range interactions are relatively rare and nonspecific, whereas some “polar” side chains form hydrogen bonds from the polar groups while occasionally forming hydrophobic contacts with the remainder of the chain.
Interaction of amylose and other α‐glucans with hydrophobic fluorescent probe (2‐p‐toluidinylnaphthalene‐6‐sulfonate)
Authors: Nakatani Hiroshi , Shibata Ken‐Ichiro , Kondo Hitoshi , Hiromi Keitaro
Published: May 2005

Fluorescence of 2‐p‐toluidinylnaphthalene‐6‐sulfonate (TNS) was enhanced in the presence of maltooligosaccharides, amylose, and other α‐glucans. The dependence of relative TNS fluorescence intensity per glucose unit on chain length of oligosaccharides was examined. The values of binding constant and thermodynamic parameters, assuming the 1:1 complex for TNS‐amylose (number‐average degree of polymerization, DPN = 17), were determined by the fluorescence titration. The values of thermodynamic parameters for 1:1 complex formation of TNS‐α‐ and β‐cyclodextrins were also determined and compared with those of TNS‐amylose (DPN = 17). The fluorescence intensity of TNS in the presence of amylose (DPN = 600) decreased by the action of glucoamylase and taka‐amylase A. The fluorescence of TNS‐amylose (DPN = 17) system increased with the increased ionic strength. In the presence of pullulan, TNS fluorescence was also enhanced and decreased by the action of pullulanase. Amylopectin enhanced TNS fluorescence rather more strongly than amylose (DPN = 17) at the same concentration. In the presence of dextran, the fluorescence of TNS was scarcely enhanced. The degree of fluorescence enhancement of TNS in the presence of α‐glucans seems to reflect the structures of α‐glucans in solution, since TNS fluorescence is enhanced in the hydrophobic environment or by the disturbance of free intramolecular rotation.
Sedimentation studies of giant DNA molecules present in unsheared whole‐cell lysates of D. melanogaster cells
Authors: Appleby David W. , Rall S. C. , Hearst John E.
Published: May 2005

We have used band sedimentation in shallow density gradients of CsCl in the preparative centrifuge to analyze the distribution of sedimentation coefficients present in tritium labelled DNA from D. melanogaster cells. The cells were lysed according to the method of Kavenoff and Zimm to preserve very high molecular weight DNA. Sedimentation measurements have been carried out as a function of speed of centrifugation. The resulting distribution functions have been interpreted with the aid of the Zimm‐Schumaker equation for the speed dependence of the sedimentation coefficient of very high molecular weight DNA. Low‐speed centrifugation (3000 rpm) indicates that DNA molecules from the lysate are evenly distributed over values of S20,w from 0 to 514S. This distribution is very sensitive to changes in speed of centrifugation and is transformed into a bimodal distribution at 12,080 rpm. Analysis of this transformation allows us to postulate that perhaps 55% of the DNA in the lysate may have molecular weights in excess of 40 × 109 g/mol. Some of these molecules may also possess a variety of configurations including partially replicated branched structures.
Catalyses by polymer complexes. VII. Enhanced esterolytic reactivity of polymer‐coenzyme A, glutathione complexes
Authors: Shinkai Seiji , Kunitake Toyoki
Published: May 2005

The association of coenzyme A(CoASH) and glutathione (GSH) with the water‐soluble polymers and their esterolytic reactivities were evaluated through the reaction with p‐nitrophenyl acetate in the presence of cationic polymer micelles: partially laurylated poly(2‐ethyl‐1‐vinylimidazole) and poly(4‐vinylpyridine). The polymer micelles with high lauryl‐group content (more than 12 mol%) markedly accelerated the reaction at very low concentrations of the polymer. Other polymers with no or small lauryl‐group content only slightly enhanced the association and the reaction rate. From the rate‐polymer concentration profiles, the association constants (K) and the rate constants for thiol coenzymes bound to the polymer (k′a,bound) were determined: for polymers with more than 12 mol % lauryl‐group content, KCoASH = 1110–2270 M−1, KGSH = 170–503M−1, k′a,bound at pH 8.65 = 142–341M−1 sec−1. k′a,bound were 20–340 times larger than that observed in the absence of the polymer. The logarithm of k′a,bound was found to be correlated well with the polymer hydrophobicity, indicating that the hydrophobic environment of the polymer activated the bound thiol anions. On the other hand, the polymer hydrophobicity did not correlate with the association constant.
Electrostatic effects in divalent ion binding to tRNA
Authors: Leroy J. L. , Guéron M.
Published: May 2005

The binding of Mn2+ to yeast tRNAPhe at 25°C is measured by esr, and found to depend strongly on the concentration of monovalent cations, showing the importance of electrostatic effects. In low sodium (<15mM/l.), the affinity is high and the Scatchard plots are distinctly curved. In high sodium (>50mM/l.), the affinity and the curvature are reduced. In a limited range of sodium concentrations (15–30mM/l.), the folding of tRNA which is induced by the divalent ions results in cooperative binding, leading to upwards convexity of the Scatchard plot. An electrostatic model is developed, based on a single type of binding site which we take to be the phosphates, with a binding constant for Mn2+ in the range of that found for ApA, 10 l./M. We compute the change in the binding constant due to the electrostatic potential of the distant charges (other phosphates and counterions), using a single set of parameters for all sodium concentrations. The model predicts that the plots in low sodium are curved, and a good fit to the experimental results is obtained: it is therefore not legitimate or necessary to interpret these results in terms of two types of binding sites. In high salt, the model gives plots that are only slightly curved, corresponding to weaker electrostatic effects. This shows that a search for sites with a special binding mode should be done in high salt. The computed plots are in good agreement with the data, except for slight differences concerning the first bound ions, which give a possible indication in favor of special binding. Given the observation of one special site for Mg2+ at 4°C in high sodium [Stein, A. & Crothers, D. M. (1976) Biochemistry 15, 157–160] in E. coli tRNAfMet, we have measured the binding of Mn2+ at lower temperature. At 12°C, in both yeast tRNAPhe and E. coli tRNAfMet, the plots clearly indicate special binding. A site found in high sodium is on a very different footing from the four to six so‐called strong sites unduly derived from low‐salt binding plots.
Vibrational modes of A‐DNA, B‐DNA, and A‐RNA backbones: An application of a green‐function refinement procedure
Authors: Lu K. C. , Prohofsky E. W. , Van Zandt L. L.
Published: May 2005

Normal vibrational analysis was carried out for DNA molecules in both A and B conformations as well as for A‐RNA. A simplified backbone model was examined and expanded to include the backbone phosphate‐group and the ribose ring. We applied the new force‐constant refinement procedure discussed in the preceeding paper [Van Zandt, L. L., Lu, K.‐C. & Prohofsky, E. W. (1977) Biopolymers, 16, 2481–90] to fit some observed frequencies in the Raman spectra for all three nucleic acids with the same set of force constants. The results indicate that the observed frequency shift can be attributed to the conformational change solely. We ignored the second‐order differences in force constants for the different geometries. The agreement between the observed and calculated frequencies derived from the final refined set of force constants is good and apparently justifies this assumption. Two modes previously assigned to the symmetric diester O‐P‐O stretch and the symmetric dioxy O‥P‥O stretch are actually fitted. They are mainly backbone phosphate‐group modes. The refined ribose‐ring force‐constants were transferred to the calculation of the vibrational spectrum of tetrahydrofuran. The overall agreement is again good. We discuss these calculations and the resulting normal modes. We also discuss the application of the Green‐function refinement scheme and several strategies adopted to bias the convergence of the procedure.
The use of light‐scattering and turbidity measurements to study the kinetics of extensively aggregating proteins: αs‐Casein
Authors: Parker T. G. , Dalgleish D. G.
Published: May 2005

The Ca2+‐induced association of αs‐casein has been studied using the methods of stopped‐flow light‐scattering and stopped‐flow turbidity. The methods used are described, and the analysis of the results to give plots of Mw against time in the time‐scale 0–15 sec is demonstrated. The validity of the method is discussed, and it is shown to be applicable to the association kinetics of aggregating proteins whose molecular‐weight averages lie between 107 and 109. The method was used to study the association of bovine αs‐casein in the presence of Ca2+, and results obtained are briefly discussed in terms of possible association mechanisms, and a mechanism for the overall reaction of αs‐casein with Ca2+, and the subsequent precipitation of the caseinate is proposed.
Calculation and use of protein‐derived conformation‐related spectra for the estimate of the secondary structure of proteins from their infrared spectra
Authors: Eckert K. , Grosse R. , Malur J. , Repke K. R. H.
Published: May 2005

This paper probes the calculation of conformation‐related basis spectra from infrared spectra (amide I′band) of reference proteins of known conformational composition and, with their aid, the computation of conformations from the amide I′ band of globular proteins using in both approaches a least‐squares, curve‐fitting computer program for the analysis of the spectra. The following results were obtained. The infrared basis spectra for the α‐helix conformation, the β‐(antiparallel‐chain pleated sheet) conformation and the ρ‐conformation were calculated and their physical reality was substantiated. The basis spectra were shown to be similar when the absorption contributions of the side chains of amino acids were either neglected or taken into account (uncorrected or corrected basis spectra). The mutual correlation of the basis spectra, quantified by the roots of the diagonal elements of the inverse matrix, was found to be low enough only for the β‐conformation to allow a statistically reliable estimate of the β‐conformation content of proteins. The comparison of the percentages of the β‐conformation derived from x‐ray structural analysis or calculated from infrared spectra showed the suitability of the basis spectra for the rough estimate of the β‐conformation percentages of proteins. The results were not significantly different when using the uncorrected or corrected basis spectra.
Investigation of the flexibility of DNA using transient electric birefringence
Authors: Hagerman Paul J.
Published: May 2005

In the preceding article, a Monte Carlo analysis was presented which provides a quantitative numerical relationship between the rotational diffusion coefficients, as measured by the decay of optical anisotropy following an electric field pulse, and the flexibility (persistence length) of short, wormlike chains. In the present article, the results of the foregoing analysis are applied to the observed rates of decay of birefringence for a series of sequenced DNA fragments ranging in size from 104 to 910 base pairs. Under the conditions used in this study, the DNA fragments exist as native, duplex molecules. Furthermore, conditions are defined in which the observed relaxation times are not dependent on DNA concentration, field strength, or the duration of the pulse. It is pointed out that the ionic atmosphere associated with a wormlike polyion does not exert any significant (direct) influence on the rotational diffusion of the polyion and, therefore, that the rotational relaxation times are a true measure of the configurations of the DNA molecules in solution. Moreover, excluded‐volume effects are shown not to be significant for the moderately short molecules employed in this study. The major conclusion of this study is that there is no strong ionic strength dependence of the persistence length for ionic strengths above 1 mM and that the persistence length, under conditions where electrostatic contributions are negligible, is approximately 500 Å. For ionic strengths significantly lower than 1 mM, electrostatic contributions to the stiffness of DNA become significant.
Rates of structural fluctuations of lysozyme in the range of thermal unfolding transition
Authors: Segawa Shin‐Ichi , Nakayama Masaaki , Sakane Masamichi
Published: May 2005

The hydrogen–deuterium exchange reaction for the tryptophan residues in lysozyme have been followed in 4.5M LiBr at pH 7.2 in the temperature range of the unfolding transition by measuring the transmittance change at 293 nm. The exchange reaction proceeded in three phases at low temperature for native protein. The first and the second phases were ascribed to the H‐D exchange reactions of three relatively exposed tryptophan residues on the molecular surface. The third phase corresponded to the H‐D exchange reaction of the three tryptophan residues buried in the interior of the molecule. The H‐D exchange reaction proceeded in two phases near the melting temperature and in a single phase at high temperature, where almost all molecules are unfolded. The H‐D exchange of three tryptophan residues buried in folded molecules was caused by fluctuation between the folded and unfolded structure of the protein molecule. The rates of such a fluctuation were determined from the rates of the exchange reaction at various temperatures. These rates agreed very well with those determined from the temperature‐jump method. This means that a protein molecule in solution fluctuates between the N‐ and D‐states at every temperature within the transition region, where the N‐form is the tightly folded native structure and the D‐form the randomly coiled chain. From measurements of thermal unfolding of ester‐108‐lysozyme and the binding constant of (NAG)3 to ester‐108‐lysozyme, it was found that almost all cross‐linked molecules are in the folded state near 50°C and pH 7.2 in 4.5M LiBr, where intact molecules are unfolded. We also studied the H‐D exchange reaction of ester‐108‐lysozyme. In the temperature region of 43–50°C, about 70% of the exchangeable tryptophan residues of ester‐108‐lysozyme were exchanged within 1 s immediately after the mixing of D2O, in spite of the fact that almost all molecules are in the folded state. This was considered the premelting of the surface of a corss‐linked molecule.
Totally synthetic crystalline ribonuclease A
Authors: Yajima Haruaki , Fujii Nobutaka
Published: May 2005

Improved chemical synthesis of bovine pancreatic ribonuclease (RNase) A was achieved by applying a new deprotecting procedure with trifluoromethanesulfonic acid–thioanisole in combination with a modified air‐oxidation procedure with glutathione for the disulfide formation. After purifications by affinity chromatography, followed by ion‐exchange chromatography, a protein with the full enzymatic activity was obtained and subsequently crystallized from aqueous ethanol according to Kunitz. A totally synthetic enzyme with full RNase A activity was thus obtained in a crystalline form for the first time.
On the location of β‐turns
Authors: Kopple Kenneth D.
Published: May 2005

β‐Turns are a common feature of cyclic peptides, but judging from recent x‐ray and solution studies of cyclic hexapeptides it is not always possible to predict in advance the type of turn and the position of the turns in the sequence. Two or more backbone conformations containing β‐turns may be of comparable energy and in rapid solvent‐ and temperature‐dependent equilibrium in solution. The use of differential relaxation effects produced by a nitroxyl radical to locate β‐turns with only minor perturbation of such equilibria is noted. Examination of the effect of a nitroxyl on the N‐H resonances of the decapeptide hormone luteinizing hormone releasing hormone supports a dominant conformation with a β‐turn at Gly6‐Leu7. Although this turn is probably part of the biologically active conformation, it is not obvious in the more active [D‐Ala6] analog.
Nmr analyses of conformations of linear peptides in solution
Authors: Miyazawa Tatsuo , Higashijima Tsutomu
Published: May 2005

Important aspects in detailed nmr analyses of the conformations of linear peptides are discussed using enkephalin and the α‐mating factor of Saccharomyces cerevisiae as examples. The cationic, dipolar, and anionic forms in dimethyl sulfoxide solution may be identified by ir analyses. Because of the electrostatic interaction between the N‐ and C‐terminal groups, the dipolar form of enkephalin takes the folded conformation, as well as extended conformation(s), in dimethyl sulfoxide solution. Such conformational equilibrium is responsible for anomalous temperature dependences and solvent‐composition dependences of the amide and Cα proton chemical shifts. Active analogs, enkephalinamide and enkephalinol, take extended conformation(s) in solution. These opioid peptides probably take a specific active conformation upon binding with a receptor. For the α‐mating factor and active peptide analogs in aqueous solution, a folded conformation with two βturn structures is responsible for the biological activity.
Conformation and activity in angiotensin II peptides
Authors: Fermandjian Serge , Piriou François , Sakarellos Constantin , Lintner Karl , Khosla Mahesh C. , Smeby Robert R. , Bumpus F. Merlin
Published: May 2005

Angiotensin II and its competitive inhibitor [Sar1, Ile8]‐angiotensin II, as well as several analogs of these two compounds specifically chosen for their well‐defined pharmacological properties, were studied by circular dichroism and nuclear magnetic resonance methods at various pH values in aqueous solution and in d6‐dimethylsulfoxide. The results were compared with their biological activities. This allowed us to establish relationships between conformation and pressor activity, explaining most of the properties of angiotensin II, its inhibitor, and the analogs successively substituted in positions 3 and 5.
Excluded volume effects on the rate of renaturation of DNA
Authors: Wetmur James G.
Published: May 2005

The rate of renaturation of T2 DNA hits been investigated by using complementary DNA strands of different length. The length of the shorter strand ranged from 0.02 to 1.0 times the length of the longer strand. An excluded volume theory is developed to include this type of reaction as well as the DNA–RNA hybridization reaction. Experimental and theoretical rates of renaturation of DNA are found to be in agreement. For the cases studied, the rate was never greater than twice that observed for short strands of the same length renaturing with themselves. The products of renaturation reactions are also considered.
Calorimetric measurement of enthalpy change in the isothermal helix–coil transition of poly‐L‐lysine in aqueous solution
Authors: Chou Peter Y. , Scheraga Harold A.
Published: May 2005

The heat ΔH° for converting an uncharged lysine residue from a coil to an α‐helical state in poly‐L‐lysine in 0.1N KCl has been determined calorimetrically to be −1200 cal/mole at both 15°C and 25°C. Essentially the same value has been obtained for the conversion of an uncharged residue from a coil to a β‐pleated sheet state. Titration data provided information about the state of charge of the polymer in the calorimetric experiments, and optical rotatory dispersion data about its conformation. In order to compute ΔH°, the observed Calorimetric heat was corrected for the heat of breaking the sample cell, the heal of dilution of HCl, the heat of neutralization of OH− ion, and the heat of ionization of the ε‐amino group in the random coil. The latter was obtained from similar Calorimetric measurements on poly‐D,L‐lysine, which was shown to be a good model for the random coil form of poly‐L‐lysine. The measured transition heat was ∼0.7 cal., which is only 7% of the total heat liberated when a 40 ml solution of 0.25% w/v poly‐L‐lysine is brought, from pH 11 to pH 7; nevertheless it could be determined with a precision of ±8%. The conformation of poly‐L‐lysine at pH 11 appears to be completely helical at 15°C, but a mixture of 90% α‐helix, 5% β form, and 5% coil at 25°C. Since ΔH° ∼ 0 for the α ⇌ β conversion, the polymer behaves like one of 95% α‐helix and 5% coil in the calorimeter at 25°C. At neutral pH, poly‐L‐lysine is an extended coil, like poly‐D,L‐lysine.