Evidence is presented that insulin antibodies produced by strain 2 guinea pigs bind preferentially to antigenic determinants related to the N-terminal B chain, N-terminal A chain, and lysine at position B-29. Strain 13 antibodies appear to bind preferentially to antigenic determinants involving the C-terminal A and C-terminal B chains. It appears that the C-terminal A chain and C-terminal B chain, in close proximity to each other, form a center rich in aromatic amino acids. This center appears to be directly related to the antigenic determinants with which the strain 13 antibodies combine, but in addition, is also important for the conformation of distant antigenic determinants. Insulin derivatives which have the greatest distortion of antigenic determinants (desoctopeptide, iodinated, and triconjugated fluorescent insulin) also have negligible biologic insulin activity. Desalamine-desasparagine insulin and diconjugated fluorescent insulin which by their immunologic reactivity appear to have distortions of antigen determinants of an intermediate degree have trace amounts of biologic activity. The insulin derivative with the least distortion of antigenic determinants (monoconjugated fluorescent insulin) had more biologic insulin activity than any of the derivatives studied.
Physical measurements involving circular dichroism and ultra centrifugal analysis confirm the immunologic observations concerning the conformations of insulin and the insulin derivatives.
These studies show that any change in conformation results in a proportionate decrease of biologic activity of insulin. Therefore, the molecular mechanism of insulin action probably involves a complementary fit with a site either on or with insulin responsive cells.
