The Tie That Binds: YOKED hCG AND DISULFIDE-LINKED SUBUNITS(1)

DISULFIDE-LINKED SUBUNITS(1)

A number of single-chain hCG agonists have been expressed and characterized, including 0(l-145)-a(l-92), 0(l-123)-a(l-92), and 0-ct(l-92), as well as several with a Ser-Gly spacer—0-(Ser-Gly)5-a(l-92), 0(1-1 ll)-(Ser-Gly)6-a, and 0(l-lO2)-(Ser-Gly)4-a. Also, various unnatural disulfide-linked subunits obtained by replacing two amino acid residues located in close proximity in the structure of HF-treated hCG, one in a and one in (3, have been reported: a-Gln5/(3-Arg8, a-Met29/(3-Met41, ct-Arg35/(B-Ala35, a-Tyr37/p-Ile38, and a-Lys51/0-Asp”. Several of these intersubunit disulfide-linked hCGs exhibit increased thermal stability relative to wild-type hCG, as does the single-chain p(l-145)-a(l-92).

The two forms, p(l-145)-a(l-92) and |3(l-123)-a(l-92), exhibit binding affinities to LHR and signaling parameters comparable to those of wild-type hCG. The fused truncated a and (3 chains with a (Ser-Gly)x spacer are also biologically active, although the binding and signaling potencies are somewhat less than the values obtained with wild-type hCG. Of the five hCG disulfide-linked heterodimers investigated, the a-Met29—>Cys/p-Met41 —>Cys mutant failed to express and the a-Lys51—»Cys/|J-Asp99—>Cys mutant exhibited only minimal potency; the other three disulfide-linked mutants were characterized by potencies only slightly less than that of wild-type hCG.

The results of competitive binding and cAMP stimulation in MA-10 cells of the full-length hCG(3-a yoked hormone expressed in baculovirus-infected insect cells are shown in Figure 3, a and b, respectively, with wild-type hCG, also expressed in baculovirus-infected insect cells, and urinary hCG included as positive controls. The apparent binding affinities of the expressed yoked hormone and heterodimer to LHR are essentially equivalent, and both are greater than that of urinary hCG. flovent inhaler
Fig3The tie that binds
FIG. 3. a) Competitive binding of [125l]hCG to transformed murine Leydig MA-10 cells by baculovirus-expressed yoked hCG with baculovirus-expressed wild-type hCG and urinary hCG included as controls, b) Cyclic AMP production in MA-10 cells in response to the same three forms of hCG. Yoked hCG (open circles), wild-type hCG (solid squares), and urinary hCG (open triangles). (Data from.)

This entry was posted in Gonadotropin and tagged Biologically Active Single-Chain Human, Chorionic Gonadotropins, Protein.