Rat uterine progesterone receptor analyzed by (/sup 3/H)R5020 photoaffinity labeling: evidence that the A and B subunits are not equimolar

Journal Article · Wed Apr 01 04:00:00 EST 1987 · Endocrinology; (United States)

OSTI ID:6526370

Ilenchuk, T T; Walters, M R

The hormone-binding components of the rat uterine progesterone receptor were investigated by the methods of (/sup 3/H)R5020 photoaffinity labeling and sodium dodecyl sulfatepolyacrylamide gel electrophoresis analysis. Two specifically labeled peaks were observed at mol wt of 85,600 +/- 1,200 and 109,600 +/- 1,200 (n = 31), resembling the A and B progesterone receptor components previously described in other systems. However, in contrast to the equimolar ratio reported in other systems, the level of subunit A observed was consistently greater than that of B (A/B ratio = 3.2 +/- 0.3; n = 31). The unusual A/B ratio prompted a complete validation of the photolabeling procedure in this system. Although the levels of specific binding increased, there was no change in the A/B ratio with varying (/sup 3/H)R5020 concentrations (5-80 nM) or with time of UV exposure (0.5 min to 3 h). Although adsorption to hydroxylapatite indicated that specific (/sup 3/H)R5020 binding was reduced by 72.0 +/- 6.4% within 5 min of UV exposure, relabeling the irradiated preparations with (/sup 3/H)R5020 resulted in little change in specific (/sup 3/H)R5020 binding. TLC analysis of (/sup 3/H)R5020 (Rf = 0.48 +/- 0.01; n = 4) after irradiation demonstrated rapid photolysis resulting in a 94.3 +/- 2.5% (n = 3) loss of authentic (/sup 3/H)R5020 within 5 min. After photolysis, at least two new tritiated products were recovered with Rf values of 0.20 +/- 0.03 and 0.72 +/- 0.02. Analysis by adsorption to hydroxylapatite indicated that the photolysis products competed for specific (/sup 3/H)R5020-binding sites in cytosol with only 10-fold lower relative binding activity than authentic R5020. Thus, these compounds probably account for the increase in specific photolabeling of the A and B peaks achieved when UV exposure is prolonged from 5 to 30 min. (Abstract Truncated)

• Research Organization: Tulane Univ. School of Medicine, New Orleans, LA
• OSTI ID: 6526370
• Journal Information: Endocrinology; (United States), Journal Name: Endocrinology; (United States) Vol. 4; ISSN ENDOA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ADSORPTION
ANIMALS
BIOCHEMICAL REACTION KINETICS
BODY
CHEMICAL REACTIONS
DECOMPOSITION
ELECTROMAGNETIC RADIATION
ELECTROPHORESIS
FEMALE GENITALS
HORMONES
ISOTOPE APPLICATIONS
KETONES
KINETICS
LABELLED COMPOUNDS
MAMMALS
MOLECULAR STRUCTURE
MOLECULAR WEIGHT
ORGANIC COMPOUNDS
ORGANS
PHOTOCHEMICAL REACTIONS
PHOTOLYSIS
PREGNANES
PROGESTERONE
RADIATIONS
RADIORECEPTOR ASSAY
RATS
REACTION KINETICS
RODENTS
SORPTION
STEROID HORMONES
STEROIDS
TRACER TECHNIQUES
TRITIUM COMPOUNDS
ULTRAVIOLET RADIATION
UTERUS
VERTEBRATES