Identification of amino-acids in the alpha-subunit first and third loops that are crucial for the heterospecific follicle-stimulating hormone activity of equid luteinizing hormone/choriogonadotropin.

The research identified specific amino acids within the alpha-subunit that play a crucial role in the variations of follicle-stimulating hormone (FSH) activity between horse (e) and donkey (dk) luteinizing hormone/choriogonadotropin (LH/CG), supporting the concept that even minute differences in hormone composition can lead to significant variations in biological activity.

Research Methodology

• The research investigated the FSH activity of eLH and dkLH. Notably, the FSH/LH ratio is ten times higher for eLH than for dkLH.
• The team created chimeric alpha-subunits containing the first 63 amino acids of the porcine (p) and the last 33 amino acids of the donkey alpha-subunit, and likewise the inverse.
• Porcine-specific amino acids were introduced by mutagenesis at set positions in donkey alpha-subunit.
• The different alpha-subunits were co-transfected in COS-7 cells with beta-eLH, beta-dkLH, and beta-eFSH.
• The bioactivity of the FSH and LH in the dimers were evaluated using two heterologous in vitro bioassays.

Research Findings

• The chimeric alpha-subunits showed FSH activity when co-expressed with beta-eLH while none was exhibited with beta-dkLH.
• The mutated hybrids displayed no FSH activity but high LH activity when expressed with beta-dkLH.
• A mutation at position 78 in alpha-dk did not affect FSH bioactivity when co-expressed with beta-eFSH.

Conclusions and Implications

• Amino acids in the first two-thirds and the last third of the equid LH alpha-subunits contribute significantly to their heterologous biospecificity. Ile alpha78 influences this to a large extent, equivalent to the beta102-103 residues; however, its effect is irrelevant to the FSH specificity of eFSH.
• The findings provide a valuable contribution to understanding the factors that govern the specific activity of these hormones and the fundamental biological differences between equid species.