Studies of the Specific Role of the Subunits of Choriogonadotropin for Biological, Immunological and Physical Properties of the Hormone. Digestion of Choriogonadotropin and Its Isolated Subunits with Serine Carboxypeptidase

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Abstract
The residues 90-92 can be split off from the C-terminal region of the isolated .alpha.-subunit of choriogonadotropin (CG, residues 88-92: -Tyr-Tyr-His-Lys-Ser-OH) by [yeast] serine carboxypeptidase (des-Lys91,Ser92-.alpha.-subunit; des-(90-92)-.alpha.-subunit). When CG is digested by serine carboxypeptidase only the residues 143-145 (-Leu-Pro-Gln-OH) form the C-terminus of the .beta.-subunit are released (des-(143-145)-CG). Depending on the pH conditions, Gln 145 and the residues 143-145, respectively, are liberated by digestion of the isolated .beta.-subunit (des-Gln145-.beta.-subunit and des-(143-145)-.beta.-subunit, respectively). The C-termini of both the isolated subunits and those in CG are probably arranged on the surface of the molecules. The biological activity of des-(143-145)-CG is not significantly decreased. The immunological activity, however, is reduced when measured by complement fixation. In comparison to the native hormone, a 4-fold amount of des-(143-145)-CG has to be applied to obtain highest complement fixation. The conformation of des-(143-145)-CG does not seem to differ from that of the native hormone, when estimated both by CD [circular dichroism] measurements and by Ans[1-anilino-8-napthalenesulfonate]-CG fluorescence. The respective determinant seems to partly depend on the sequence of the C-terminal region of the .beta.-subunit of the hormone; complement fixation does not seem to be significantly affected, when the des-(143-145)-.beta.-subunit is compared with the native .beta.-subunit using an antiserum against the native .beta.-subunit. This C-terminal determinant is possibly more immunogenic at the hormone than at the isolated .beta.-subunit. The biological activity of recombined CG in vivo as well as in vitro is markedly reduced when Ser 92 is removed from the C-terminus of the .alpha.-subunit (des-Ser92,Lys91-.alpha.-+ native .beta.-subunit: 36% residual activity in vivo). Biological activity is lost when the residues 88-90 are removed by digestion of the des-Ser92,Lys91-.alpha.-subunit with [bovine pancreatic] carboxypeptidase A [EC 3.4.17.1]. Recombination products between a modified .alpha.- and the native .beta.-subunit show a reduced Ans-CG fluorescence (des-Lys91,-Ser92-.alpha.-+ native .beta.-subunit: 52%; des-(88-92)-.alpha.-+ native .beta.-subunit: 23%). The Ans-induced aggregation of CG also takes place in those recombination products which display a low Ans-CG fluorescence, indicating that the reduction is probably not caused by a portion of the molecules losing their binding sites for Ans. The diminished Ans-CG fluorescence seems to signal small conformational changes. The CD spectra of the native and the des-(90-92)-.alpha.-subunit, do not significantly differ. The release of amino acids from the C-terminus of the .alpha.-subunit causes a disturbance of the interaction between the subunits. This seems to prevent an effective conformational change of the .beta.-subunit which probably is a prerequisite for the binding of the hormone to the receptors of Leydig cells.

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