Conformational properties of human immunoglobulin G subclasses analysis by temperature-perturbation and solvent-perturbation spectroscopy

Abstract

Conformational properties of human myeloma immunoglobulins G belonging to four subclasses (IgG1 Van, IgG2 Kom, IgG3 Pla, IgG4 Ang), and also Fab, Fc and pFc′ fragments derived from IgG1 Van, IgG2 Kom and IgG3 Pla have been studied by temperature-perturbation and solvent-perturbation spectroscopy. It has been shown that the immunoglobulins studied practically do not differ in the number of tyrosine and tryptophan residues exposed to different solvent perturbants (saccharose, glycerol, dimethylsulfoxide). The same regularity is observed for isolated Fab and Fc fragments. At the same time, the immunoglobulins compared and their proteolytic fragments significantly differ in the number of aromatic chromophores perturbed by temperature. These data indicate that immunoglobulins of different subclasses and their subunits have a different rigidity of structure in relation to thermal perturbation. The Fc subunits of IgG1 are characterized by the lowest rigidity of structure of internal hydrophobic cores of domains (characterized by the rigidity of the microenvironment of tryptophan residues), as compared with the Fc subunits of IgG2 and IgG3. In the case of IgG1 and IgG2, these differences seem to be brought about by a different rigidity of structure of C H2 domains, since thermal-perturbation spectra of the pFc′ fragments of these subclasses practically coincide. The total number of chromophores exposed to different solvent perturbants in the isolated Fab and Fc fragments practically coincides with the number of exposed chromophores in intact immunoglobulins. Similar coincidence is observed for the tryptophan residues perturbed by temperature. These data indicate that neither the conformation of surface sites nor the conformation of internal hydrophobic cores of domains significantly changes on isolation of Fab and Fc fragments. At the same time, many more tyrosine residues are perturbed by temperature in the intact immunoglobulin G1 Van than in the corresponding sum of isolated Fab and Fc fragments, while for IgG2 Kom, which has the same length of hinge region, these values practically coincide. This fact can be explained by the greater temperature dependence of motions of subunits in IgG1 Van as compared with IgG2 Kom, and as a result of this by the higher mutual temperature-dependent influence of subunits on their internal structure (on interdomain interactions).