Precision interferometry as a new method for studying the conformational state of protein and its interaction with a solvent

A new method for studying the conformational state of a protein and its interaction with a solvent has been developed. The method is based on measuring the refractive index of a protein solution using a precision laser interferometer with an error of 10$^{-7}$. The interferometer was calibrated on the basis of the known dependences of the refractive index change with a change in water temperature, and the refractive index change of a NaCl solution with a change in concentration. Using the interferometer, the refractive index changes were first recorded in the course of proteolysis (during pepsin-catalyzed hydrolysis, an increase in the refractive index of solutions of bovine serum albumin and egg lysozyme by 9 $\times$ 10$^{-6}$ and 2.4 $\times$ 10$^{-6}$, respectively) and denaturation (an increase in the refractive index by 4.5 $\times$ 10$^{-5}$ during the reaction of egg lysozyme with guanidine hydrochloride and dithiothreitol). Based on direct interferometric measurements, we have disproved the widespread assumption that the refractive index of protein solutions is determined only by concentration of the protein and its amino acid composition and does not depend on the state of protein fragmentation. The increased accuracy of the interferometer makes it possible to study the refractive index increment $(dn/dc)$ at low concentrations of dissolved compounds ($\ll$ 1%), as well as to examine the processes leading to changes in the conformation of macromolecules in solution.