The time course of recombination of human chorionic gonadotropin subunits studied with immunological methods, circular-dichroic measurements and bioassay.

Abstract

The restitution of immunological properties of native human chorionic gonadotropin by recombination of the subunits, α and β, was studied by immunological methods (qualitative and two‐dimensional immunoelectrophoresis and two immunodiffusion methods). Recombination of the subunits at different incubation times was carried out with three different mixture ratios. The bulk of the recombined hormone is formed during the first hour. The immunological behaviour of recombined hormone corresponds largely to that of the starting material used for the preparation of the subunits. In addition to the recombined hormone material a further recombination product appears which can be detected neither in the subunit preparations nor in the native hormone. It seems to be formed in high concentrations immediately after the beginning of incubation and to decrease rapidly thereafter. With an excess of β subunit in the recombination mixture, however, it does not disappear completely. The second product possesses antigenic determinants different from those of the recombined hormone. The circular dichroic spectra of the recombination mixtures display variations in dichroic absorption during the course of recombination, as well as a third maximum. With increasing incubation time, the third maximum decreases and is shifted to the far ultraviolet region, where it finally overlaps with the extremum at 195 nm.The biological activity of the reconstituted material reaches as much as 75% of the starting material.