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A biologically active single chain human chorionic gonadotropin analog with altered receptor binding properties
1Department of Biochemistry and Molecular Biology, University of Georgia, Athens 30602-7229, USA. narayan@bchiris.bmb.uga.edu
Endocrinology
|December 30, 1999
Summary
New single-chain human chorionic gonadotropin (hCG) analogs were engineered. One analog, YhCG3, shows lower receptor binding affinity but equal efficacy, suggesting altered structural requirements for hCG receptor activation.
Area of Science:
- Endocrinology and Reproductive Biology
- Molecular and Cellular Biology
- Hormone Receptor Interactions
Background:
- Human chorionic gonadotropin (hCG) is a glycoprotein hormone essential for pregnancy, composed of common alpha and unique beta subunits.
- Engineered single-chain hCG analogs (e.g., YhCG1) have been created by fusing subunits to study receptor binding and activation.
- Previous designs (N-beta-alpha-C) were based on the presumed importance of hCGbeta N-terminus and alpha-subunit C-terminus for receptor interaction.
Purpose of the Study:
- To construct and characterize a novel single-chain hCG analog (YhCG3) with an inverted subunit order (N-alpha-CTP-beta-C).
- To compare the receptor binding affinity and biological activity of YhCG3 with a previously engineered analog (YhCG1).
- To investigate the role of free subunit termini in hCG receptor interaction and activation.
Main Methods:
- Construction of a new single-chain hCG analog, YhCG3, by fusing the alpha-subunit C-terminus to the hCGbeta N-terminus via a C-terminal peptide (CTP) linker.
- Assessment of receptor binding affinity of YhCG3 compared to YhCG1.
- Measurement of intracellular cAMP levels in transfected cells stimulated by YhCG3 and YhCG1.
- Evaluation of YhCG3's ability to form a covalently linked, biologically active complex with its receptor.
Main Results:
- YhCG3 exhibited a 25- to 30-fold lower binding affinity to the LH receptor compared to YhCG1.
- Despite lower affinity, YhCG3 stimulated intracellular cAMP production to an extent comparable to YhCG1.
- YhCG3 formed a covalently linked, biologically active complex with the LH receptor, leading to elevated basal cAMP levels.
Conclusions:
- The free N- and C-termini of hCGbeta and the alpha-subunit, respectively, are not critical for receptor binding and activation.
- YhCG3 represents a conformationally distinct single-chain hCG analog that is more efficacious in activating the LH receptor than YhCG1.
- These findings provide new insights into the structural determinants of hCG-receptor interactions and hormone efficacy.