The antibodies against the computationally designed mimic of the glycoprotein hormone receptor transmembrane domain provide insights into receptor activation and suppress the constitutively activated receptor Mutants

Majumdar, Ritankar and Railkar, Reema and Dighe, Rajan R (2012) The antibodies against the computationally designed mimic of the glycoprotein hormone receptor transmembrane domain provide insights into receptor activation and suppress the constitutively activated receptor Mutants. In: Journal of Biological Chemistry, 287 (41). pp. 34514-34532.

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Abstract

The exoloops of glycoprotein hormone receptors (GpHRs) transduce the signal generated by the ligand-ectodomain interactions to the transmembrane helices either through direct hormonal contact and/or by modulating the interdomain interactions between the hinge region (HinR) and the transmembrane domain (TMD). The ligand-induced conformational alterations in the HinRs and the interhelical loops of luteinizing hormone receptor/follicle stimulating hormone receptor/thyroid stimulating hormone receptor were mapped using exoloop-specific antibodies generated against a mini-TMD protein designed to mimic the native exoloop conformations that were created by joining the thyroid stimulating hormone receptor exoloops constrained through helical tethers and library-derived linkers. The antibody against the mini-TMD specifically recognized all three GpHRs and inhibited the basal and hormone-stimulated cAMP production without affecting hormone binding. Interestingly, binding of the antibody to all three receptors was abolished by prior incubation of the receptors with the respective hormones, suggesting that the exoloops are buried in the hormone-receptor complexes. The antibody also suppressed the high basal activities of gain-of-function mutations in the HinRs, exoloops, and TMDs such as those involved in precocious puberty and thyroid toxic adenomas. Using the antibody and point/deletion/chimeric receptor mutants, we demonstrate that changes in the HinR-exoloop interactions play an important role in receptor activation. Computational analysis suggests that the mini-TMD antibodies act by conformationally locking the transmembrane helices by means of restraining the exoloops and the juxta-membrane regions. Using GpHRs as a model, we describe a novel computational approach of generating soluble TMD mimics that can be used to explain the role of exoloops during receptor activation and their interplay with TMDs.

Item Type:	Journal Article
Publication:	Journal of Biological Chemistry
Publisher:	American Society for Biochemistry and Molecular Biology
Additional Information:	Copyright of this article belongs to American Society for Biochemistry and Molecular Biology.
Department/Centre:	Division of Biological Sciences > Molecular Reproduction, Development & Genetics
Date Deposited:	20 Feb 2013 09:03
Last Modified:	20 Feb 2013 09:03
URI:	http://eprints.iisc.ac.in/id/eprint/45383

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