Aquametric submicro determination of water content of biopolymer preparations and its application in deriving hydration isotherms.
A rapid aquametric submicro method is proposed for determining 10-100,mug of bound water in 20-60 mug amounts of biopolymers. This method has been applied in deriving isotherms of water-vapour adsorption by biopolymer preparations (proteins, tRNA), in a dynamic mode requiring only 5 mg of the substance. The BET equation was used to determine the effective capacity (h) of a monolayer in the course of sorption (h --> ) and desorption (h <-- ), corresponding to accessible primary hydration sites on the biopolymer molecule surface. As has been established for proteins with known spatial structure, omega-amide groups of Asn and Gln residues and ion-pair forming groups do not participate in the formation of the BET-monolayer during sorption. Such an interpretation of the isotherms underlies the estimation of the number of surface and screened polar groups in the molecules of biopolymers with a spatial structure not yet established from amino-acid analysis and hydration isotherms. The maximum hydration (H(s)) of globular proteins is much less pronounced than that of tRNA, which can be explained by an irregular arrangement of sterically separated primary hydration sites which do not form a matrix for distribution and ordering of water at great distances. The deficiency of sorption sites because of sterically inaccessible groups is a trait common to biopolymers in general. Therefore, hydration isotherms may characterize certain aspects of the macromolecular structure, such as compactness, degree of screening of the polar groups, regularity of arrangement of the primary hydration sites, and so on.
Sherman
F B
FB
N.D. Zilinsky Institute of Organic Chemistry, U.S.S.R. Academy of Sciences, 47 Leninsky Prospect, Moscow, 117334, U.S.S.R.
Khurgin
Y I
YI