Unfortunately, we do not have accurate measures of receptor density for the yoked human hCG-LHR complex expressed on HEK 293 cells, and thus it is impossible to estimate a potency based, for example, on the basal cAMP expression levels and numbers of complex per cell. Moreover, there is no information on turnover, internalization rates, desensitization, etc. of the yoked hCG-LHR complex.
The yoked hCG-receptor complex should prove very useful for gonadotropin and receptor structure-function studies, as well as studies aimed at elucidating the mechanism of transmembrane signaling. Some caution is in order in interpreting the constitutive activation of LHR by a yoked ligand. It is assumed, but has not been proven, that the conformations of the ligand and receptor are similar, when present as a single-chain construct, to those adopted when the natural heterodimer binds to and activates the wild-type receptor. While this is a plausible assumption, it nonetheless remains to be verified experimentally. buy asthma inhalers
Since the yoked hCG-LHR complex exhibited such striking intrinsic activity, we asked whether the individual subunits could function as agonists if expressed in singlechain forms with LHR. The answer is yes. Figure 5 shows that the basal cAMP levels are elevated in HEK 293 cells expressing yoked human a-LHR and yoked hCG(3-LHR relative to mock-transfected cells and cells transfected with LHR; the (3 subunit, however, acts as a much more potent agonist than the a subunit when linked to LHR via CTP and a Factor Xa cleavage site.
FIG. 5. Basal cAMP production in mock-transfected HEK 293 cells (MT) and cells transfected with cDNAs to wild-type LHR (LHR), yoked human a-LHR (YaR), and yoked hCGp-LHR (YfJR).