Structural Features of γ-Immunoglobulin, Antibody, and Their Fragments. Circular Dichroism Studies

Renata E. Cathou, Anthony Kulczycki, Edgar Haber

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The far-ultraviolet circular dichroism of rabbit antidinitrophenyl antibody is the sum of separate contributions from the Fab and Fc fragments. Analysis of the circular dichroism of Fab revealed the presence of at least seven bands, located at 217 (-), 232 (+), 242 (-), 265 (+), 275 (+), 285 (+), and 291 mμ (+), all of which were found to be sensitive to the conformational state of the protein. In particular, the latter four optically active transitions, which had not been previously observed in optical rotatory dispersion studies, disappeared when the protein was exposed to 4 m guanidine hydrochloride for 5 hr; these bands are due to tyrosine, tryptophan, and/or cystine in asymmetric environments. The negative band at 217 mμ is probably due to β structure. The positive band at 232 mμ has been tentatively assigned to un-ionized tyrosine residues on the basis of molar ellipticity and alteration on raising the pH to 12. The ellipticity of the negative band at 242 mμ and the effect on it of exposing Fab to guanidine hydrochloride suggest that random coil may be the source of this band, although cystines may also make contributions. The circular dichroism of nonspecific rabbit γ-immunoglobulin Fab showed bands at the same wavelengths as those of antidinitrophenyl Fab but the molar ellipticities of all bands were smaller. The circular dichroism of antidinitrophenyl Fab in the presence of a threefold molar excess é-dinitrophenyllysine was the same as that of the protein in the absence of ligand indicating that no conformational changes detectable by circular dichroism had occurred on formation of the antibody-hapten complex.

Original languageEnglish
Pages (from-to)3958-3964
Number of pages7
Issue number11
StatePublished - Nov 1 1968


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