Partial molar volumes of the amino acid side-chains of proteins in aqueous solution: some comments on their estimation using partial molar volumes of amino acids and small peptides.

The partial molar volumes of some amino acid side-chains were determined in two recent studies [Makhatadze, G.I., et al. (1990) Biopolymers 30, 1001, and Reading, J.F. & Hedwig, G.R. (1990) J. Chem. Soc. Faraday Trans. 86, 3117] using partial molar volume data, V02, in aqueous solution at 25 degrees C for some peptides of sequence Gly-X-Gly, where X is an amino acid. These side-chain partial molar volumes are critically compared with those obtained using V02 data for amino acids. It is concluded that side-chain partial molar volumes calculated using V02 data for the tripeptides are better estimates of side-chain partial molar volumes in proteins than are those determined using V02 data for amino acids.     

Partial molar volumes of polypeptides and their constituent groups in aqueous solution over a broad temperature range

The partial molar volumes of various compounds that model protein constituent groups, such as tripeptides (Gly-X-Gly, where X = Gly, Ala, Val, Leu, Ile, Pro, Met, His, Ser), homopeptides (Glyn, n = 3,4,5), and simple organic analogues of amino acid side chains (methanol, acetamide, propanamide, acetic acid, propanoic acid, n-butanamine, n-butanamine nitrate, n-propylguanidine nitrate, 4-methylphenol), have been determined in aqueous solution with a vibrational densimeter in the temperature range of 5-85 degrees C. The partial molar volumes of amino acid side chains and the peptide unit were estimated from the data obtained. Assuming additivity of component groups, the partial molar volumes of polypeptide chains of several proteins over a broad temperature range were calculated. The partial molar volume functions of four proteins (myoglobin, cytochrome C, ribonuclease A, lysozyme) were compared with those determined experimentally for the unfolded and native forms of these proteins. It has been shown that the average deviation of the calculated functions from the experimental ones does not exceed 3% over the temperature range studied.     

Partial molar volumes of some alpha-amino acids in aqueous sodium acetate solutions at 308.15 K

The apparent molar volumes V(2,phi) have been determined for glycine, DL-alpha-alanine, DL-alpha-amino-n-butyric acid, DL-valine and DL-leucine in aqueous solutions of 0.5, 1.0, 1.5 and 2.0 mol kg(-1) sodium acetate by density measurements at 308.15 K. These data have been used to derive the infinite dilution apparent molar volumes V(0)(2,phi) for the amino acids in aqueous sodium acetate solutions and the standard volumes of transfer, Delta(t)V(0), of the amino acids from water to aqueous sodium acetate solutions. It has been observed that both V(0)(2,phi) and Delta(t)V(0) vary linearly with increasing number of carbon atoms in the alkyl chain of the amino acids. These linear correlations have been utilized to estimate the contributions of the charged end groups (NH(3)(+), COO(-)), CH(2) group and other alkyl chains of the amino acids to V(0)(2,phi) and Delta(t)V(0). The results show that V(0)(2,phi) values for (NH(3)(+), COO(-)) groups increase with sodium acetate concentration, and those for CH(2) are almost constant over the studied sodium acetate concentration range. The transfer volume increases and the hydration number of the amino acids decreases with increasing electrolyte concentrations. These facts indicate that strong interactions occur between the ions of sodium acetate and the charged centers of the amino acids. The volumetric interaction parameters of the amino acids with sodium acetate were calculated in water. The pair interaction parameters are found to be positive and decreased with increasing alkyl chain length of the amino acids, suggesting that sodium acetate has a stronger dehydration effect on amino acids which have longer hydrophobic alkyl chains. These phenomena are discussed by means of the co-sphere overlap model.     

A new set of peptide-based group heat capacities for use in protein stability calculations.

The partial molar heat capacities of the tripeptides of the sequence glycyl-X-glycine, where X is one of the amino acids leucine, threonine, glutamine, phenylalanine, histidine, cysteine, proline, glutamic acid or arginine, and of the two tetrapeptides tetraglycine and glycyltryptophanylglycylglycine in aqueous solution over the temperature range 10-100 degrees C have been determined using high sensitivity scanning microcalorimetry. These results were used to derive the partial molar heat capacities of the various amino acid side-chains. This report completes our programme to derive reliable side-chain heat capacities for all 20 amino acids of proteins over a wide temperature range using the tripeptides Gly-X-Gly as realistic model compounds. Included in the study is a summary of the partial molar heat capacities of all 20 amino acid side-chains. These results, along with the heat capacity of the peptide backbone group, were used to calculate the partial molar heat capacities of some oligopeptides and of the random coil form of some unfolded proteins in water. The calculated heat capacities of the proteins obtained using this new set of heat capacities for the constituent groups are consistent with the heat capacities of the denatured state determined experimentally.     

Partial molar heat capacities of the peptides glycylglycylglycine, glycyl-L-alanylglycine and glycyl-DL-threonylglycine in aqueous solution over the temperature range 50 to 125 degrees C

Partial molar heat capacities of the three tripeptides glycylglycylglycine, glycyl-L-alanylglycine and glycyl-DL-threonylglycine in aqueous solution at the temperatures 50, 75, 100 and 125 degrees C have been determined using differential flow calorimetry. The results have been used to estimate the contributions to the partial molar heat capacities of peptides of the alanyl and threonyl side-chains. These side-chain contributions are compared with those reported in the literature.     

The partial molar heat capacity and volume of the peptide backbone group of proteins in aqueous solution

The partial molar heat capacities have been determined for the series of peptides alanyl(glycyl)(x)glycine, x=1-3, and for the compounds N-acetylglycinamide and N-acetyl glycylglycinamide in aqueous solution over the temperature range 10-100 degrees C using high sensitivity scanning microcalorimetry. The partial molar volumes for these compounds have also been determined over the temperature range from 10 to 90 degrees C using a scanning densimetric method. The results were used to derive the partial molar heat capacities and volumes of the glycyl group at temperatures in the range 10-100 degrees C. The results obtained are critically compared with literature results derived using heat capacity and volume data for some oligoglycines.     

Partial molar volumes and adiabatic compressibilities of unfolded protein states

We determined the partial molar volumes, V degrees , and adiabatic compressibilities, K degrees (S), of N-acetyl amino acids with neutralized carboxyl termini, N-acetyl amino acid amides, and N-acetyl amino acid methylamides between 18 and 55 degrees C. The individual compounds in the three classes have been selected so as to collectively cover the 20 naturally occurring amino acid side chains. We interpret our experimental results in terms of the volumetric contributions and hydration properties of individual amino acid side chains and their constituent atomic groups. We also conducted pH-dependent densimetric and acoustic measurements to determine changes in volume and compressibility accompanying protonation of the aspartic acid, glutamic acid, histidine, lysine, and arginine side chains. We use our resulting data to develop an additive scheme for calculating the partial molar (specific) volume and adiabatic compressibility of fully extended polypeptide chains as a function of pH and temperature. We discuss the differences and similarities between our proposed scheme and the reported additive approaches. We compare our calculated volumetric characteristics of the fully extended conformations of apocytochrome c and apomyoglobin with the experimental values measured in water (for apocytochrome c) or acidic pH (for apomyoglobin). At these respective experimental conditions, the two proteins are unfolded. However, the comparison between the calculated and experimental volumetric characteristics suggests that neither apocytochrome c nor apomyoglobin are fully unfolded and retain a sizeable core of solvent-inaccessible groups.     

Viscosity B-coefficients and standard partial molar volumes of amino acids, and their roles in interpreting the protein (enzyme) stabilization

This review systematically surveys the viscosity B-coefficients and standard partial molar volumes of amino acids at various temperatures as these data are quite important for interpreting the hydration and other properties of peptides and proteins. The effect of organic solutes and various ions on the viscometric and volumetric properties of amino acids has also been discussed in terms of their kosmotropic ('structure-making') effects on the hydration of amino acids. The comparison of these effects on the amino acid hydration enables us to have a better understanding of the influence of organic solute and salt on the protein stabilization. In addition, the viscometric and volumetric behaviors of amino acid ions (cations and anions) are also summarized because these ions have recently been incorporated as part of novel ionic liquids, which have wide applications in biocatalysis and protein stabilization.     

Thermodynamics of some nucleic acid bases and nucleosides in water, and their transfer to aqueous glucose and sucrose solutions at 298.15 K

The partial molar heat capacities and volumes of some of the constituents of nucleic acids have been determined in water and 1 molal aqueous glucose and sucrose solutions in order to elucidate the nature of interactions occurring between various nucleic acid bases, nucleosides and the sugar (glucose and sucrose) molecules. The results have been explained in terms of the contributions from hydrophobic interactions, hydrophilic interactions and the hydrogen bonding between the solute and solvent molecules. The results have also been compared with those of amino acids and peptides in aqueous glucose and sucrose solutions.     

Aqueous solutions containing amino acids and peptides. Part 20. Volumetric behavior of some terminally substituted amino acids and peptides at 298.15 K

The densities at 298.15 K of aqueous solutions containing some terminally substituted amino acids and peptides containing the glycyl, L -and D -alanyl, L -leucyl, sarcosyl, and L -prolyl residues have been dertermined and standard state partial molar volumes and volumetric pairwise virial coefficients obtained. It is shown that the partial molar volumes can be represented using group volume contributions, but this approach is only approximate, and significant effects of N-terminal substitution and sequence dependence are observed. The volumetric virial coefficients for the amino acid amides have been expressed using a group-additivity approach, and the results obtained indicate that the dominant contributions come from peptide group interactions with other peptide groups and with hydrophobic groups. There is also some evidence of both sequence and chiral effects on the volumetric virial coefficients for proline-containing dipeptides.     

Isentropic and isothermal compressibilities of the backbone glycyl group of proteins in aqueous solution

The partial molar isentropic compressibilities at infinite dilution, K(S,2)(o), have been determined for the peptides serylglycine, serylglycylglycine and serylglycylglycylglycine in aqueous solution at 25 degrees C. The partial molar volumes at infinite dilution, V(2)(o), have also been determined for these peptides in aqueous solution at the temperatures 15, 30 and 40 degrees C. These results, along with those obtained previously at 25 degrees C, were used to derive the partial molar exansibilities, E(2)(o), of the peptides at 25 degrees C, which in turn were used to convert the isentropic compressibilities into the partial molar isothermal compressibilities at infinite dilution, K(T,2)(o). These K(S,2)(o) and K(T,2)(o) results were used to obtain the partial molar compressibilities of the glycyl group CH(2)CONH at 25 degrees C. The results are compared with those obtained using data for other series of peptides of sequence ala(gly)(n), n=1-4, and (gly)(n), n=2-5.     

Hydration of nucleic bases in dilute aqueous solutions. Apparent molar adiabatic and isothermal compressibilities, apparent molar volumes and their temperature slopes at 25 degrees C

The concentration increment of the ultrasound velocity has been measured with an accuracy of +/- 0.03 cm/s in dilute aqueous solutions of a variety of nucleic bases and their derivatives in the concentration range 0.5-1.5 mg/g H2O at temperatures of 15-35 degrees C. A new method for the precise measurement of ultrasound velocity in small volumes of liquids has been used. The values of the apparent molar adiabatic compressibilities plus the corresponding temperature slopes, apparent molar volumes with their temperature slopes, and apparent molar isothermal compressibilities at infinite dilution have been obtained. The regularities describing the signs of these values and their dependence on the chemical structure of the solute have been revealed. It is shown that these regularities can be described as a consequence of partial 'normalization' of some of the properties of water around the bases, namely, weaker structural contribution to compressibility, less negative temperature slope of compressibility and less negative structural contribution to the coefficient of thermal expansion of water.     

The partial molar volumes of anesthetics in lipid bilayers

The excess volumes of mixing of benzyl alcohol, halothane, and methoxyflurane in water and in suspensions of several lipid bilayers have been determined at 25 degrees C using a novel excess volume dilatometer. The excess volumes of mixing in water were all found to be negative, whereas in lipid suspensions they were all more positive than those in water alone. From known partition coefficients the partial molar volumes of these three solutes in the lipid bilayers were calculated. These values were all close to the molar volumes of the pure anesthetics, as was a value determined for halothane in olive oil. Halothane was studied in dipalmitoylphosphatidylcholine below its phase transition, and was found to exhibit a much larger excess volume than in any other system we studied. The potency of these three anesthetics was determined in tadpoles. It was calculated that at equi-anesthetic doses these three agents caused an expansion in egg lecithin/cholesterol (2:1) bilayers of 0.21 +/- 0.015%. This result is consistent with the hypothesis that general anesthetics act by expanding membranes.     

Partial molar volumes of some α-amino acids in aqueous sodium acetate solutions at 308.15 K

The apparent molar volumes V_2,φ have been determined for glycine, -α-alanine, -α-amino-n-butyric acid, -valine and -leucine in aqueous solutions of 0.5, 1.0, 1.5 and 2.0 mol kg⁻¹ sodium acetate by density measurements at 308.15 K. These data have been used to derive the infinite dilution apparent molar volumes V⁰_2,φ for the amino acids in aqueous sodium acetate solutions and the standard volumes of transfer, Δ_tV⁰, of the amino acids from water to aqueous sodium acetate solutions. It has been observed that both V⁰_2,φ and Δ_tV⁰ vary linearly with increasing number of carbon atoms in the alkyl chain of the amino acids. These linear correlations have been utilized to estimate the contributions of the charged end groups (NH₃⁺,COO⁻), CH₂ group and other alkyl chains of the amino acids to V⁰_2,φ and Δ_tV⁰. The results show that V⁰_2,φ values for (NH₃⁺,COO⁻) groups increase with sodium acetate concentration, and those for CH₂ are almost constant over the studied sodium acetate concentration range. The transfer volume increases and the hydration number of the amino acids decreases with increasing electrolyte concentrations. These facts indicate that strong interactions occur between the ions of sodium acetate and the charged centers of the amino acids. The volumetric interaction parameters of the amino acids with sodium acetate were calculated in water. The pair interaction parameters are found to be positive and decreased with increasing alkyl chain length of the amino acids, suggesting that sodium acetate has a stronger dehydration effect on amino acids which have longer hydrophobic alkyl chains. These phenomena are discussed by means of the cosphere overlap model.     

Effect of calcium chloride on the conformation of proteins. Thermodynamic studies of some model compounds

Partial molar heat capacities (CP2 degrees) and volumes (V2 degrees) for some amino acids and peptides were measured in 1 M aqueous calcium chloride solutions at 298.15 degrees K using a Picker flow microcalorimeter and an oscillating-tube digital density meter. Using the data for these amino acids and peptides in water, the corresponding partial molar heat capacities of transfer (CP2,tr degree) and volumes (V2,tr degree) from water to 1 M aqueous calcium chloride were deduced. These thermodynamic parameters are significantly positive, indicating that strong interactions occur between the ions of calcium chloride and the charged centres of these amino acids and peptides. A comparison has been made with a similar transfer of these compounds to sodium chloride solutions. The thermodynamic parameters of the transfer of peptide group (-CONH) are much more positive in calcium chloride than in sodium chloride solutions. The implication of this result for the ability of calcium chloride to act as a stronger destabilizer of protein conformation is discussed.     

Partial molar volumes of mRNA 5' cap analogues

Partial molar volumes in aqueous solution of eleven selected 7-methylguanine cap-analogues and their guanine counterparts were determined by means of density measurements. Hydrophobicity of the investigated compounds regarding their structural features was analysed within the framework of the solute-solvent interaction model, based on the relative density of the molecular solvation shell.     

Ionic partial molar volumes of density gradient salts at finite concentrations.

A general method is presented for estimating concentration-dependent partial molar volumes for individual ions of heavy density gradient salts used in the isopycnic ultracentrifugation of charged macromolecules.     

Apparent molar volumes of some amino acids and peptides in aqueous urea solutions

Densities of solutions of several α-amino acids and peptides in 3 and 6m aqueous urea solvents have been determined at 298.15 K. These data have been used to evaluate the infinite-dilution apparent molar volumes of the solutes and the volume changes due to transfer (V ) of the α-amino acids and peptides at infinite dilution from water to aqueous urea solutions. The sign and magnitude of the V values have been rationalized in the framework of Friedman's cosphere-overlap model. The V values for the glycyl group (? CH₂CONH? ) and alkyl side chains have been estimated.     

E.s.r. of spin-trapped radicals in aqueous solutions of amino acids. Reactions of the hydroxyl radical.

The radicals produced by reactions of hydroxyl radicals with amino acids in aqueous solutions have been investigated. Hydroxyl radicals were formed by U.V.-photolysis of hydrogen peroxide and the short-lived amino acid radicals were spin-trapped by tert-nitrosobutane and identified by electron spin resonance spectroscopy. Nineteen amino acids were studied, and several radicals were identified which have not been observed previously by other methods. Only side-chain radicals were identified for alanine, threonine, aspartic acid, asparagine, lysine, phenylalanine, tyrosine, proline and hydroxyproline; whereas for glycine the C(2) carbon radical was spin-trapped. Both C(2) carbon radicals and side-chain radicals were assigned to valine, leucine, isoleucine, serine, glutamic acid, glutamine, arginine and methionine.     

A convenient general method for synthesis of N alpha- or N omega-dithiasuccinoyl (Dts) amino acids and dipeptides: application of polyethylene glycol as a carrier for functional purification

N alpha-Dithiasuccinoyl (Dts) amino acids needed for solid-phase peptide synthesis have been prepared in good yields and excellent purities by a new method that exploits the solubility properties of polyethylene glycol (PEG; bifunctional with average molecular weight 2000 was found to be optimal). Suitably side-chain protected amino acid derivatives are first reacted with a polymeric xanthate, following which the free alpha-carboxyl is blocked by silylation and the Dts heterocycle is elaborated in the same pot by reaction with chlorocarbonylsulfenyl chloride. Upon aqueous work-up, the polymeric carrier removes any urethane blocked amino acids which arise during the process. Exaggerated conditions were explored to prove the power of this functional purification approach, and mechanisms of formation of polymer-bound urethanes are proposed and supported by solution model studies. The preparation and characterization of the companion N-(iso-propyldithio)carbonyl derivative of proline is also presented.