EQUINE ANTIHAPTEN ANTIBODY . MOLECULAR WEIGHTS OF SUBUNITS OF EQUINE IMMUNOGLOBULINS

Three independent methods have been used to determine the molecular weights of the heavyand light-polypeptide chain subunits of equine γGab-, γGC-, and γT-immunoglobulins. Extensively reduced and alkylated proteins were filtered through standard columns of Sephadex G-100 or G-200 in 8 M urea-0.05 m propionic acid. Subunit molecular weights were obtained from the linear elution volume, Ve, vs. logarithm molecular weight relationship defined for each column with rabbit γG-globulin heavy and light chains and horse heart cytochrome c. Molecular weights also were determined by equilibrium sedimentation in 6 m guanidine hydrochloride. Partial specific volumes, ̅v, were estimated from the amino acid composition of the polypeptide chains and reduced by a factor of 0.015 ml/g to eliminate the effect of preferential binding of guanidine hydrochloride. The recovery of protein in the light fractions, obtained upon Sephadex G-200 gel filtration of extensively reduced and alkylated proteins in 0.04 m sodium decyl or dodecyl sulfate, was used as a third parameter to estimate subunit molecular weights. The three methods furnished molecular weight values which were in substantial agreement. The equine immunoglobulin heavy and light chains had molecular weights of 52,200-53,900 and 22,300-23,100, respectively. A molecular weight of 53,100-54,900 was obtained for the heavy chain of a human γA myeloma protein. © 1969, American Chemical Society. All rights reserved.