Thyrotropin receptors in normal human thyroid. Nonclassical binding kinetics not explained by the negative...

Area of Science:

• Endocrinology
• Molecular Biology
• Biochemistry

Background:

• Thyrotropin (TSH) is crucial for thyroid function, binding to specific receptors on thyroid cells.
• Understanding TSH binding kinetics and affinity is essential for elucidating thyroid regulation.
• Previous studies suggested complex binding interactions, potentially involving cooperativity or multiple receptor subtypes.

Purpose of the Study:

• To analyze the equilibrium binding of iodinated TSH (125I-TSH) to human thyroid preparations.
• To differentiate between receptor binding and non-specific binding under various conditions.
• To investigate the phenomenon of negative cooperativity in TSH binding.

Main Methods:

• Saturation analysis of 125I-TSH binding to normal human thyroid preparations.
• Scatchard plot analysis under physiological and non-physiological buffer conditions (pH, ionic strength).
• Computer analysis to resolve binding site components and kinetic experiments on 125I-TSH dissociation.

Main Results:

• Linear Scatchard plots were observed under non-physiological conditions (pH 6.0 or 20 mM Tris), indicating a single binding affinity (approx. 10(-8) M).
• Non-linear Scatchard plots under standard conditions (pH 7.4, 40 mM Tris) revealed at least two binding sites: high-affinity (approx. 10(-10) M) and low-affinity.
• Non-target tissues showed binding only under non-physiological conditions, with similar affinity but lower capacity than normal thyroid.

Conclusions:

• 125I-TSH binds to heterogeneous sites in human thyroid tissue, with high-affinity sites likely representing the TSH receptor.
• The observed negative cooperativity phenomenon appears to be an artifact of non-physiological conditions and low-affinity binding, possibly to gangliosides.
• Standard physiological conditions are necessary for accurate characterization of high-affinity TSH receptor binding.