1796
The Journal of Physical Chemistry, Vol. 83, No. 13, 1979
A. P.
Sarvazyan, D. P. Kharakoz, and P. Hemmes
Ultrasonic Investigation of the pH-Dependent Solute-Solvent Interactions in Aqueous Solutions of Amino Acids and Proteins A. P. Sarvazyan,* D. P. Kharakor, Institute of Bio/ogica/ Physics, Academy of Sciences of the USSR, Puschino, Moscow Region, USSR 142292
and Paul Hemmes” Department of Chemistry, Rutgers, The State University of New Jersey, Newark, New Jersey 07102 (Received February 5, 1979)
Ultrasonic velocity titrations have been performed in aqueous solution on glycine, alanine, and histidine. All systems were studied in the pH range 5-13. Histidine shows two inflections corresponding to it’s two pK values in this pH range. The more acidic process shows a symmetrical behavior about pH = pK. All amino acids studied showed an asymmetric behavior if the pH is above 9. This asymmetry is due to a relaxational velocity dispersion which occurs for the deprotonation reaction. This effect has its maximum value at a pH near 11. The titration curve can be corrected for this effect by studies of the absorption of sound. When corrected, the titration curves can be interpreted in terms of hydrating changes which accompany the reaction. It is found that the NH3+ group on glycine is the most hydrated. The other amino acids show lower hydration due to steric effects. These results were extended to the protein, metmyoglobin. This substance shows three inflections in the titration curve. These correspond to titration of histidine residues, free NH3+ groups (including a relaxational contribution), and a velocity change due to denaturation of the protein by base. Semiquantitative agreement is found between the theoretical and experimental titration curves.
The state of ionization of the surface of biomolecules and related hydration and proton transfer process of these sites are a significant part of the interactions of these molecules with their environment. Ultrasonic absorption spectroscopy has long played a leading role in research involving the kinetics and thermodynamics of proton transfer reactions since the relaxation times of these processes lie in the region of time covered by this experimental technique ( T < s). Another aspect of molecular acoustics, the measurement of ultrasonic velocities, has played only a small role in the investigation of biologically significant molecules. However, such measurements can provide important information about the inter- and intramolecular interactions. 1-3 The development of precision methods for measurements of the propagation velocity of ultrasound in small sample volumes by means of resonator t e c h n i q u e P expands the capabilities of ultrasonic investigation of compressibilities and hydration of macromolecules in solution. The objective of this work is to show the capability of this method in the investigation of the state of ionization of biopolymers by using for examples the velocity titration of metmyoglobin and amino acids in neutral and basic solution. The sound velocity, U , is related to the coefficient of adiabatic compressibility, P, and the solution density, p, by the equation
P=
(1) Therefore any change in solution structure accompanied by a change in compressibility is reflected in a change in velocity. The parameters which characterize the solute in solution are the apparent molar compressibility @K 3 (Pv - PoVo)/n and the related quantity the relative increment of sound velocity A = AU/(UoC)where Po, Vo,and Uoare the compressibility, volume, and velocity of solvent and P and V, the corresponding quantities for a volume of solution containing n moles of solute. The quantity AU is the difference between velocity of ultrasound in solution and in the solvent. (PV)F1
The Velocity of Ultrasound and Hydration The relationship between GK and A is given, for an aqueous solution, by the approximate expression valid for dilute solutions where AU
