Extreme enthalpy‒entropy compensation in the dimerization of small solutes in aqueous solution

This communication summarizes findings from the earliest encounters with extreme enthalpy‒entropy compensation, a phenomenon first detected in the 1950s by a reappraisal of isopiestic and calorimetric measurements on aqueous urea solutions in terms of solute self-association. Because concurrent studies of carboxylic acid association were confined to measurement of the equilibrium constant by conductance, IR spectrophotometry or potentiometric titration measurements, temperature-independence of the dimerization constant was mistakenly taken to signify a value of zero for Δ H o instead of (ΔH o ‒ TΔS o ). In those studies of small-solute self-association the extreme enthalpy‒entropy compensation was reflecting the action of water as a reactant whose hydroxyl groups were competing for the solute carbonyl involved in self-association. Such action gives rise to a positive temperature dependence of Δ H o that could well be operating in concert with that responsible for the commonly observed negative dependence for protein‒ligand interactions exhibiting extreme enthalpy‒entropy compensation, where the solvent contribution to the energetics reflects changes in the extent of ordered water structure in hydrophobic environments

Concentration dependence of translational diffusion coefficients for globular proteins

This investigation examines published results of traditional diffusion experiments on ovalbumin and bovine serum albumin to determine the extent to which assumed concentration independence of the translational diffusion coefficient is a reasonable approximation in the analysis of boundary spreading in sedimentation velocity experiments on proteins. Although significant positive concentration dependence of the diffusion coefficient (D) for both proteins is predicted by current theories, none has been detected in these experimental diffusion studies performed under the constraints of constant temperature and solvent chemical potential (those also pertinent to sedimentation velocity). Instead, the results are better described by the relatively minor concentration dependence predicted by considering solution viscosity to be an additional source of D–c dependence. Inasmuch as the predicted variation in D for solutions with concentrations below 10 mg mL-1 is within the uncertainty of experimental estimates, these findings support use of the approximate solution of the Lamm equation developed by Fujita for the quantitative analysis of boundary spreading in sedimentation velocity experiments on proteins

Quantifying the concentration dependence of sedimentation coefficients for globular macromolecules: a continuing age-old problem

This retrospective investigation has established that the early theoretical attempts to directly incorporate the consequences of radial dilution into expressions for variation of the sedimentation coefficient as a function of the loading concentration in sedimentation velocity experiments require concentration distributions exhibiting far greater precision than that achieved by the optical systems of past and current analytical ultracentrifuges. In terms of current methods of sedimentation coefficient measurement, until such improvement is made, the simplest procedure for quantifying linear s-c dependence (or linear concentration dependence of 1/s) for dilute systems therefore entails consideration of the sedimentation coefficient obtained by standard c(s), g*(s) or G(s) analysis) as an average parameter (s¯) that pertains to the corresponding mean plateau concentration (following radial dilution) (c¯) over the range of sedimentation velocity distributions used for the determination of s¯. The relation of this with current descriptions of the concentration dependence of the sedimentation and translational diffusion coefficients is considered, together with a suggestion for the necessary improvement in the optical system

Thermodynamics of semi-specific ligand recognition: the binding of dipeptides to the E.coli dipeptide binding protein DppA

This investigation of the temperature dependence of DppA interactions with a subset of three dipeptides (AA. AF and FA) by isothermal titration calorimetry has revealed the negative heat capacity (ΔCpo) that is a characteristic of hydrophobic interactions. The observation of enthalpy–entropy compensation is interpreted in terms of the increased structuring of water molecules trapped in a hydrophobic environment, the enthalpic energy gain from which is automatically countered by the entropy decrease associated with consequent loss of water structure flexibility. Specificity for dipeptides stems from appropriate spacing of designated DppA aspartate and arginine residues for electrostatic interaction with the terminal amino and carboxyl groups of a dipeptide, after which the binding pocket closes to become completely isolated from the aqueous environment. Any differences in chemical reactivity of the dipeptide sidechains are thereby modulated by their occurrence in a hydrophobic environment where changes in the structural state of entrapped water molecules give rise to the phenomenon of enthalpy–entropy compensation. The consequent minimization of differences in the value of ΔG0 for all DppA–dipeptide interactions thus provides thermodynamic insight into the biological role of DppA as a transporter of all dipeptides across the periplasmic membrane

Oligomeric state of hypoxanthine-guanine phosphoribosyltransferase from Mycobacterium tuberculosis

Sedimentation equilibrium and size-exclusion chromatography experiments on Mycobacterium tuberculosis hypoxanthine-guanine phosphoribosyltransferase (MtHGPRT) have established the existence of this enzyme as a reversibly associating mixture of dimeric and tetrameric species in 0.1\ua0M Tris-HCl−0.012\ua0M MgCl, pH 7.4. Displacement of the equilibrium position towards the larger oligomer by phosphate signifies the probable existence of MtHGPRT as a tetramer in the biological environment. These data thus add credibility to the relevance of considering enzyme function in the light of a published tetrameric structure deduced from X-ray crystallography. Failure of 5-phospho-α-D-ribosyl-1-pyrophosphate (PRib-PP) to perturb the dimer−tetramer equilibrium position indicates the equivalence and independence of binding for this substrate (the first to bind in an ordered sequential mechanism) to the two oligomers. By virtue of the displacement of the equilibrium position towards dimer that is affected by removing MgCl from the Tris-HCl buffer, it can be concluded that divalent metal ions, as well as phosphate, can affect the oligomerization. These characteristics of MtHGPRT in solution are correlated with published crystal structures of four enzyme−ligand complexes