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The beta104-109 sequence is essential for the secretion of correctly folded single-chain beta alpha horse LH/CG and for its FSH activity.

Authors
  • Galet, Colette
  • Guillou, Florian
  • Foulon-Gauze, Florence
  • Combarnous, Yves
  • Chopineau, Maryse
Type
Published Article
Journal
Journal of Endocrinology
Publisher
Bioscientifica
Publication Date
Oct 01, 2009
Volume
203
Issue
1
Pages
167–174
Identifiers
DOI: 10.1677/JOE-09-0141
PMID: 19589909
Source
Medline
License
Unknown

Abstract

The dual LH and FSH activity of the equine LH (eLH)/equine chorionic gonadotropin (eCG) in heterologous species makes eLH/CG a good model to study structure/function relationships of gonadotropins. In order to bypass the problem of intracellular association of the heterodimer, a recombinant single-chain beta alpha eLH/CG was used to identify sequences in the beta-subunit involved in the secretion and activities of the hormone. The C-terminal region of the beta-subunit was progressively truncated. All resulting truncated single-chains were secreted in the media as detected by an anti-beta peptide antibody in reducing conditions. However, using a conformation sensitive ELISA we show that the truncated single-chains were differently recognized: deletion of the last 40 amino acids of the beta-subunit (beta109alpha eLH/CG) resulted in a 90% decrease in the recognized correctly folded hormone compared with the full-length beta alpha eLH/CG single-chain and no properly folded hormone was detected in the secretion medium when the last 46 amino acids of the beta-subunit were deleted (beta103alpha eLH/CG). We thus focused on the six amino acids sequence 104-109, which belongs to the seat-belt region. Mutation of the 104-109 sequence in alanines in the full-length beta alpha eLH/CG (beta104-109Ala alpha) led to a 50% decrease in the production of properly folded hormone in COS-7 as well as in alphaT3 pituitary cells. Moreover, the FSH activity of this mutant was decreased by 70% whereas its LH activity remained intact. These data lead us to conclude that the 104-109 region of the beta eLH/CG subunit is essential for the secretion of a fully folded beta alpha eLH/CG and for its FSH activity but not for its LH activity.

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