Characterization of the thyrotropin receptor
Scatchard analysis of the binding of (/sup 125/I)TSH to thyroid plasma membranes results in a curvilinear, concave-upward plot. This phenomenon could be indicative of several conditions, including radioligand heterogeneity, negative cooperativity, or multiple binding sites. To investigate the first of these possibilities, (/sup 125/I)TSH was purified by chromatography on Sepharose 6B. The receptor active (/sup 125/I)TSH continued to yield a curvilinear Scatchard plot in equilibrium binding analyses, indicating that this phenomenon was not the result of radioligand impurities of heterogeneity. To determine the contribution of the receptor to this complex behavior, the TSH receptor was solubilized and subjected to concanavalin A chromatography. Two populations of binding sites were recovered. The pass-through fraction contained 70% of the total sites and exhibited a linear Scatchard plot with a K/sub D/ of 67 nM, while 0.2 M methylmannoside eluted 10% of the sites which exhibited a single K/sub D/ of 0.3 nM. To characterize its structure, the TSH receptor was labeled with (/sup 125/I)TSH and cross-linked with disuccinimidyl suberate. Analysis by electrophoresis and autoradiography demonstrated the labeling of two hormone-receptor complexes with M/sub r/ of 80,000 and 68,000. These two bands were demonstrated to be TSH-specific and were present in plasma membranes from thyroid, but not from muscle or liver. Furthermore, antibodies isolated from the sera of Graves' disease patients, which inhibit the bindings of (/sup 125/I)TSH, blocked the labeling of the two complexes. When the separated high and low affinity TSH binding components were similarly analyzed, both components exhibited the 80,000 and 68,000 bands. Furthermore, the autoantibodies from Graves' disease sera inhibited the binding of (/sup 125/I)TSH to both the high and low affinity species.
- Research Organization:
- North Carolina Univ., Chapel Hill (USA)
- OSTI ID:
- 7027066
- Resource Relation:
- Other Information: Thesis (Ph. D.)
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
RECEPTORS
BIOCHEMICAL REACTION KINETICS
CHEMICAL COMPOSITION
TSH
ANTIBODIES
AUTORADIOGRAPHY
CELL MEMBRANES
CHROMATOGRAPHY
ELECTROPHORESIS
IMMUNE SERUMS
IODINE 125
LABELLED COMPOUNDS
MOLECULAR WEIGHT
THYROID
TRACER TECHNIQUES
BETA DECAY RADIOISOTOPES
BODY
CELL CONSTITUENTS
DAYS LIVING RADIOISOTOPES
ELECTRON CAPTURE RADIOISOTOPES
ENDOCRINE GLANDS
GLANDS
HORMONES
INTERMEDIATE MASS NUCLEI
IODINE ISOTOPES
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
MEMBRANE PROTEINS
MEMBRANES
NUCLEI
ODD-EVEN NUCLEI
ORGANIC COMPOUNDS
ORGANS
PEPTIDE HORMONES
PITUITARY HORMONES
PROTEINS
RADIOISOTOPES
REACTION KINETICS
SEPARATION PROCESSES
550201* - Biochemistry- Tracer Techniques