Estimation of molecular parameters of the endogenous inhibitor to progesterone--receptor binding in rat trophoblast.
This study was undertaken to investigate molecular properties of an endogenous substance which inhibits progesterone--receptor binding. The inhibitor was isolated from the cytosol of rat trophoblast by ammonium sulfate precipitation. Further purification was achieved by gel filtration and sucrose density gradient centrifugation. These techniques revealed that the inhibitor had a Stokes radius of 2.8 +/- 0.1 nm and an s of 4.9 +/- 0.2. For a protein exhibiting normal density and solvation, these parameters indicated that the inhibitor molecule has a mol. wt of 56,000 +/- 2000 g/mol and a frictional coefficient of 1.11 +/- 0.01. In order to obtain an additional independent estimate of molecular weight, the migratory pattern of the inhibitor was studied on SDS-polyacrylamide gels after identification and purification by polyacrylamide gel electrophoresis. The inhibitor-active fraction was resolved into two principle bands (Band 1 and 2) by SDS-gel electrophoresis having mol. wt of 59,000 +/- 700 g/mol and 51,000 +/- 300 g/mol (n = 6), respectively. The molecular weight thus determined was in excellent agreement with the value obtained by calculation. Thus, molecular parameters of the inhibitor indicate that it is a very symmetrical molecule of approx. 56,000 mol. wt. Characterization of the molecular properties of the inhibitor substance should facilitate future studies concerning the biological significance of this molecule and its role in progesterone--receptor binding interactions.
D000818:Animals; D002499:Centrifugation, Density Gradient; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D003600:Cytosol / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D008970:Molecular Weight; D011374:Progesterone / Q000378:metabolism; D051381:Rats; D011980:Receptors, Progesterone / Q000037:antagonists & inhibitors* / Q000378:metabolism; D014327:Trophoblasts / Q000032:analysis*
D011980:Receptors, Progesterone; D011374:Progesterone