Extracellular domain of lutropin/choriogonadotropin receptor expressed in transfected cells binds choriogonadotropin with high affinity.

Xie, Yanbo; Wang, Haiyun; Segaloff, Deborah L.

doi:10.1016/s0021-9258(18)45750-x

Cited by 224 publications

(29 citation statements)

References 33 publications

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“…This is consistent with previous observations indicating the presence of a FSH binding site in the N-terminal half of the receptor (27,28). Similarly truncated N-terminal halves of the LH/CG receptor and the TSH receptor are capable of high affinity hormone binding (17)(18)(19)24). This high affinity site is, however, incapable of activating the receptors (19).…”

Section: Resultssupporting

confidence: 92%

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Differential Roles of Exoloop 1 of the Human Follicle-stimulating Hormone Receptor in Hormone Binding and Receptor Activation

1995

Journal of Biological Chemistry

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The follicle-stimulating hormone (FSH) receptor is a member of the glycoprotein hormone receptor subfamily of the seven-transmembrane receptor superfamily. These receptors have an extracellular N-terminal half of approximately 350 amino acids and a membrane-associated C-terminal half of approximately 350 amino acids. The N-terminal halves have the high affinity hormone binding site. On the other hand, the C-terminal half of the lutropin/choriogonadotropin receptor has the receptor activation site. However, little is known about the activation site and mechanism of the FSH receptor, although the existing evidence indicates crucial differences in the activation of the FSH receptor and the lutropin/choriogonadotropin receptor. As a first step to resolve this issue, we examined the upstream juxtamembrane five amino acids, Asp405-Ile406-His407-Thr408-Lys409, of the exoloop 1. Ala scan and multi-substitutions show that the five amino acid sequence is important for both hormone binding and receptor activation to induce cAMP synthesis, despite its short length. Specifically, His407 is important for high affinity hormone binding, whereas Asp405, Thr408, and Lys409 are crucial for receptor activation. The data suggest that the five amino acids may form a turn of helix that is an extension of the transmembrane helix 2. In this helical arrangement, Asp405, Thr408, and Lys409 are grouped to form a hydrophilic face of the helix, suggesting a correlation between this arrangement and receptor activation. In addition, the diverse and differential roles of the five amino acids indicate that high affinity hormone binding and receptor activation are discernible functions. These novel observations will be helpful for understanding the activation mechanism of the FSH receptor.

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Section: Resultssupporting

confidence: 92%

“…The receptor has high and low affinity hormone contact sites. The high affinity site is in the extracellular N-terminal half (17)(18)(19), and the low affinity site is in the membrane-associated C-terminal half (20). The low affinity site alone is capable of activating the LH/CG receptor to induce hormone action (19)(20)(21)(22).…”

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confidence: 99%

Differential Roles of Exoloop 1 of the Human Follicle-stimulating Hormone Receptor in Hormone Binding and Receptor Activation

1995

Journal of Biological Chemistry

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“…LHR has been expressed in different models not exempt from drawbacks; for instance, in human embryonic kidney cells, the expressed extracellular domain shows high a nity to hCG but remains trapped within the cells [5]. Similarly, LHR expressed in baculovirus-infected insect cells is inactive and remains trapped in aggregate pools [6].…”

Section: Resultsmentioning

confidence: 99%

“…In the testis, LHR is activated by LH, also called interstitial cell-stimulating hormone, initiating the transduction process via activating cAMP, and the reaction pathways to synthesize testosterone by Leydig cells [4]. The second large domain of LHR, encoded into the lhr gene, comprises the long eleventh exon, which is distinctive of the GPCR structure, formed by seven transmembrane α-helices and a cytoplasmic tail corresponding to the carboxy-terminus [5].…”

Section: Introductionmentioning

confidence: 99%

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Expression and Purification of the Extracellular Domain of Luteinizing Hormone Receptor From Ovis Aries Testicle

Villalpando-Aguilar¹,

López-Rosas

Montero-Pardo

et al. 2021

Preprint

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The luteinizing hormone receptor (LHR) is a glycoprotein member of the G protein-coupled receptor superfamily. Physiologically, this receptor participates in corpus luteum formation and ovulation in females. In males, it acts in testosterone synthesis and spermatogenesis and is involved in some fertility disorders. RNA was extracted from Ovis aries testicles, and the corresponding cDNA was synthesized to amplify the lhr gene, termed lhr-bed here, consisting of 762 bp that encodes 273 amino acids of the extracellular domain of LHR. Thus, the lhr-bed was cloned into pJET1.2/blunt, subcloned into the pCOLD II expression vector and finally transformed into E. coli BL21 cells. Since the induced rLHR-Bed protein was found in the insoluble fraction, the purification protocol was modified as follows: induction at 25°C, denaturing conditions (8 M UREA and 0.1% CHAPS) and refolding in the column to increase solubility. The rLHR-Bed expression was corroborated by western blotting and mass spectrometry (MS) analysis. This successful method to obtain the recombinant LHR extracellular domain yields 0.2 mg/L of the protein with approximately 90% purity from a single chromatographic purification step. The present approach demonstrates the feasibility of obtaining large quantities of rLHR-Bed. This might be useful to accomplish future studies regarding the structure and functional analysis of the binding interplay with its ligand luteinizing hormone and its isoforms. Additionally, this biotechnological strategy might be used to improve and to develop new drugs for the treatment of reproductive disorders and might also be applied to species reproduction in the livestock industry.

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Molecular architecture and biorecognition processes of the cystine knot protein superfamily: part I. The glycoprotein hormones

2000

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Extracellular domain of lutropin/choriogonadotropin receptor expressed in transfected cells binds choriogonadotropin with high affinity.

Cited by 224 publications

References 33 publications

Differential Roles of Exoloop 1 of the Human Follicle-stimulating Hormone Receptor in Hormone Binding and Receptor Activation

Differential Roles of Exoloop 1 of the Human Follicle-stimulating Hormone Receptor in Hormone Binding and Receptor Activation

Expression and Purification of the Extracellular Domain of Luteinizing Hormone Receptor From Ovis Aries Testicle

Molecular architecture and biorecognition processes of the cystine knot protein superfamily: part I. The glycoprotein hormones

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