Structural and Functional Studies of the Tryptic Core of the Human Chorionic Gonadotropin β-Subunit*

Publisher Website
Google Scholar
Abstract
The .beta.-subunit of hCG was digested with trypsin to produce a modified form of the subunit for structure-function and immunological studies. After digestion of hCG.beta. with trypsin, the residual disulfide-linked core was isolated and found to be lacking the carboxy-terminal peptide (residues 115-145) and to contain bond cleavages between residues 2-3, 43-44, 74-75, and 95-96. The locations of these bond cleavages within the disulfide-bridged core were identified by isolation of the following peptides after reduction and S-carboxymethylation of the trypsin .beta.-core: .beta.1-43, .beta.3-43, .beta.44-74, .beta.44-95, .beta.75-95, and .beta.96-114. The circular dichroic spectrum of the tryptic .beta.-core over the wavelength region of about 200-320 nm was similar to that of the native subunit. In addition, the tryptic .beta.-core retained nearly full immunopotency in both polyclonal and monoclonal competitive RIAs and could combine with complementary native .alpha.-subunit. The hybrid, composed of the tryptic .beta.-core and native .alpha., was purified and displayed a molar potency of about 0.1% relative to intact hCG in both a radioreceptor assay and an adenylate cyclase assay. Thus, the hybrid retained little biological activity. Although the extensive bond cleavages in the tryptic .beta.-core did not appear to change its secondary and tertiary structure sufficiently to significantly alter the circular dichroic spectrum, the immunoreactivity, or the capability to combine with its .alpha.-subunit complement, the biological functional integrity of the tryptic .beta.-core-containing hybrid was essentially abolished. Hence, the tryptic .beta.-core provides a useful derivative for detailed structure-function studies aimed at defining the necessary determinants for subunit association, receptor binding, and subsequent biological actions.