Glycerol Decreases the Volume and Compressibility of Protein Interior

Abstract

The addition of hydrogen-bonded cosolvents to aqueous solutions of proteins is known to modify both thermodynamic and dynamic properties of the proteins in a variety of ways. Previous studies suggest that glycerol reduces the free volume and compressibility of proteins. However, there is no directly measured evidence for that. We have measured the apparent specific volume (V) and adiabatic compressibility (K) of a number of proteins, sugars, and amino acids in water and in 30% glycerol at pH 7.4 and 30 degrees C. The values of V and K in water and their changes induced by glycerol were extrapolated to the limit of infinite solute size. The main results were the following: (a) glycerol decreases V and K of proteins, but increases it for amino acids; (b) the V and K values of the protein interior in water were found to be 0.784 +/- 0.026 mL/g and (12.8 +/- 2.5) x 10(-6) mL/g x atm, where the glycerol reduces these values by 8 and 32%, respectively; (c) the coefficient of adiabatic compressibility of the structural component of proteins affected by the glycerol is estimated to be (50 +/- 10) x 10(-6) atm(-1), which is comparable to that of water. We propose that the glycerol induces a release of the so-called "lubricant" water, which maintains conformational flexibility by keeping apart neighboring segments of the polypeptide chain. This is expected to lead to the collapsing of the voids containing the water as well as to increase intramolecular bonding, which explains the observed effect.

Effect of glycerol and dimethyl sulfoxide on the phase behavior of lysozyme: Theory and experiments

“…III A 2). This is supported by observations of Priev et al 19 Their experiments indicate an increase of the hydration shell upon addition of glycerol and, at the same time, a decrease of the core volume of BPTI. Similarly, using small-angle neutron scattering, Sinibaldi et al 20 observed, upon increasing the glycerol content, a significant increase of the hydration shell of lysozyme from 0.3 nm to 0.6 nm, and a small decrease of the core volume of lysozyme of about 6%, such that the total effective protein volume increases by at least about 13%.…”

Dynamics and Structural Changes Induced by ATP Binding in SAV1866, a Bacterial ABC Exporter

“…Water has been proposed to serve as a lubricant favoring the folding/unfolding process of proteins. 72 A similar role could be conceived for water in the association/ dissociation process of proteins. These changes at the NBD-TMD interface must propagate to the TMD at the extracellular side.…”

Octameric alcohol oxidase dissociates into stable, soluble monomers upon incubation with dimethylsulfoxide

“…A wealth of data is available on the effect of glycerol on protein stability (for a review see [17]). The cosolvent glycerol is preferentially excluded from the protein-solvent interface and can therefore induce a decrease in volume and compressibility of the protein interior [13]. It has been suggested that a balance between repulsion of glycerol from hydrophobic surfaces of the protein and interaction with polar regions, is in fact the origin of the well known stabilizing effect of glycerol [18,19].…”

“…Three scans were averaged for each sample and standard deviations were derived from three separate samples. Buffer blanks were subtracted and secondary structure composition estimated using the program CONTIN [13]. Temperature scans were conducted at a wavelength of 222 nm, with scan rates of 2°C/min and accumulation times of 4 s. Melting points (T m ) were estimated as the temperature at which 50% AO is unfolded.…”