• Genomics & Informatics
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• v.19 no.1
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• pp.9.1-9.5
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• 2021

Mammalian olfactory receptors are a family of G protein-coupled receptors (GPCRs) that occupy a large part of the genome. In human genes, olfactory receptors account for more than 40% of all GPCRs. Several types of GPCR structures have been identified, but there is no single olfactory receptor whose structure has been determined experimentally to date. The aim of this study was to model the interactions between an olfactory receptor and its ligands at the molecular level to provide hints on the binding modes between the OR2W1 olfactory receptor and its agonists and inverse agonists. The results demonstrated the modes of ligand binding in a three-dimensional model of OR2W1 and showed a statistically significant difference in binding affinity to the olfactory receptor between agonists and inverse agonists.

• Biomolecules & Therapeutics
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• v.25 no.1
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• pp.57-68
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• 2017

Seven transmembrane receptors (7TMRs), also known as G protein-coupled receptors, are popular targets of drug development, particularly 7TMR systems that are activated by peptide ligands. Although many pharmaceutical drugs have been discovered via conventional bulk analysis techniques the increasing availability of structural and evolutionary data are facilitating change to rational, targeted drug design. This article discusses the appeal of neuropeptide-7TMR systems as drug targets and provides an overview of concepts in the evolution of vertebrate genomes and gene families. Subsequently, methods that use evolutionary concepts and comparative analysis techniques to aid in gene discovery, gene function identification, and novel drug design are provided along with case study examples.

• Journal of Life Science
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• v.17 no.6 s.86
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• pp.783-790
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• 2007

G protein coupled receptors (GPCRs) transmit various extracellular signals into the cells. Upon binding of the ligands, conformational changes in the extracellular and/or transmembrane (TM) domains of CPCRs were propagated into the cytoplasmic (CP) domain of the molecule leading to the activation of their cognate heterotrimeric C proteins and kinases. Constitutively active GPCR mutants causing the activation of C Protein signaling even in the absence of ligand binding are of interest for the study of activation mechanism of GPCRs. Two classes of constitutively active mutations, categorized by their effects on the salt bridge between Ell3 and K296, were found in the TM domain of rhodopsin. Opsin mutants containing combinations of the mutations were constructed to study the conformational changes required for the activation of rhodopsin. Rhodopsin chromophore regenerated with 11-cis-retinal showed a thermal stability inversely correlated with its constitutive activity. In contrast, rhodopsin mutants exhibited a binding affinity to an agonist, all-trans-retinal, in a constitutive activity-dependent manner. In order to test whether the conformational changes responsible for the activation of trans-ducin (Gt) are the same as the conformation required for the recognition of rhodopsin kinase, analysis of the mutants were carried out with phosphorylation by rhodopsin kinase. Rhodopsin mutants containing combinations of different classes of the mutations showed a strong synergistic effect on the phosphorylation of the mutants in the dark as similar to that of Gt activation. The results suggest that at least two or three kinds of segmental and independent conformational changes are required for the activation of rhodopsin and the conformational changes responsible for activating rhodopsin kinase and Gt are similar to each other.

• Molecules and Cells
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• v.41 no.2
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• pp.150-159
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• 2018

Animals use their odorant receptors to receive chemical information from the environment. Insect odorant receptors differ from the G protein-coupled odorant receptors in vertebrates and nematodes, and very little is known about their protein-protein interactions. Here, we introduce a mass spectrometric platform designed for the large-scale analysis of insect odorant receptor protein-protein interactions. Using this platform, we obtained the first Orco interactome from Drosophila melanogaster. From a total of 1,186 identified proteins, we narrowed the interaction candidates to 226, of which only two-thirds have been named. These candidates include the known olfactory proteins Or92a and Obp51a. Around 90% of the proteins having published names likely function inside the cell, and nearly half of these intracellular proteins are associated with the endomembrane system. In a basic loss-of-function electrophysiological screen, we found that the disruption of eight (i.e., Rab5, CG32795, Mpcp, Tom70, Vir-1, CG30427, Eaat1, and CG2781) of 28 randomly selected candidates affects olfactory responses in vivo. Thus, because this Orco interactome includes physiologically meaningful candidates, we anticipate that our platform will help guide further research on the molecular mechanisms of the insect odorant receptor family.

• The Korean Journal of Physiology and Pharmacology
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• v.14 no.3
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• pp.145-150
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• 2010

Adenosine (Ado) is an important mediator of the endogenous defense against ischemia-induced injury in the heart. The action of Ado is mediated by activation of G protein-gated inwardly rectifying $K^+$ (GIRK) channels. In turn, GIRK channels are inhibited by reducing phosphatidylinositol 4,5-bisphosphate ($PIP_2$) through Gq protein-coupled receptors (GqPCRs). We previously found that GIRK channels activated by acetylcholine, a muscarinic M2 acetylcholine receptor agonist, are inhibited by GqPCRs in a receptor-specific manner. However, it is not known whether GIRK channels activated by Ado signaling are also regulated by GqPCRs. Presently, this was investigated in mouse atrial myocytes using the patch clamp technique. GIRK channels were activated by $100\;{\mu}M$ Ado. When Ado was repetitively applied at intervals of 5~6 min, the amplitude of second Ado-activated GIRK currents ($I_{K(Ado)}$) was $88.3{\pm}3.7%$ of the first $I_{K(Ado)}$ in the control. Pretreatment of atrial myocytes with phenylephrine, endothelin-1, or bradykinin prior to a second application of Ado reduced the amplitude of the second $I_{K(Ado)}$ to $25.5{\pm}11.6%$, $30.5{\pm}5.6%$, and $96.0{\pm}2.7%$, respectively. The potency of $I_{K(Ado)}$ inhibition by GqPCRs was different with that observed in acetylcholine-activated GIRK currents ($I_{K(ACh)}$) (endothelin-1>phenylephrine>bradykinin). $I_{K(Ado)}$ was almost completely inhibited by $500\;{\mu}M$ of the $PIP_2$ scavenger neomycin, suggesting low $PIP_2$ affinity of $I_{K(Ado)}$. Taken together, these results suggest that the crosstalk between GqPCRs and the Ado-induced signaling pathway is receptor-specific. The differential change in $PIP_2$ affinity of GIRK channels activated by Ado and ACh may underlie, at least in part, their differential responses to GqPCR agonists.

• Proceedings of the Korean Biophysical Society Conference
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• 2003.06a
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• pp.29-29
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• 2003

Compartmentation of intracellular signaling pathways serves as an important mechanism conferring the specificity of G protein-coupled receptor (GPCR) signaling. In the heart, stimulation of $\beta$$_2$-adrenoceptor ($\beta$$_2$-AR), a prototypical GPCR, activates a tightly localized protein kinase A (PKA) signaling, which regulates substrates at cell surface membranes, bypassing cytosolic target proteins (eg, phospholamban). Although a concurrent activation of $\beta$$_2$-AR-coupled $G_{i}$ proteins has been implicated in the functional compartmentation of PKA signaling, the exact mechanism underlying the restriction of the $\beta$$_2$-AR-PKA pathway remains unclear. In the present study, we demonstrate that phosphatidylinositol 3-kinase (PI3K) plays an essential role in confining the $\beta$$_2$-AR-PKA signaling. Inhibition of PI3K with LY294002 or wortmannin enables $\beta$$_2$-AR-PKA signaling to reach intracellular substrates, as manifested by a robust increase in phosphorylation of phospholamban, and markedly enhances the receptor-mediated positive contractile and relaxant responses in cardiac myocytes. These potentiating effects of PI3K inhibitors are not accompanied by an increase in $\beta$$_2$-AR-induced cAMP formation. Blocking $G_{i}$ or $G_{$\square$$\square$}$ signaling with pertussis toxin or $\beta$ARK-ct, a peptide inhibitor of $G_{$\square$$\square$}$, completely prevents the potentiating effects induced by PI3K inhibition, indicating that the pathway responsible for the functional compartmentation of $\beta$$_2$-AR-PKA siglaling sequentially involves $G_{i}$, $G_{$\square$$\square$}$, and PI3K. Thus, PI3K constitutes a key downstream event of $\beta$$_2$-AR- $G_{i}$ signaling, which confines and negates the concurrent $\beta$$_2$-AR/Gs-mediated PKA signaling.gnaling.

• Biomedical Science Letters
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• v.15 no.4
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• pp.327-333
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• 2009

Heterotrimeric GTP binding proteins, G-proteins, mediate signal transduction generated by neurotransmitters and hormones. Among G-proteins, Go proteins are the most abundant in brain and classified as a member of Gi family. Ras-like protein in all tissues (Rit), one of the small GTPases, is a member of a Ras superfamily and identified as an important regulator of neuronal differentiation and cell transformation. Recently, we have reported that Rit functioned as a candidate downstream effector for alpha subunit of Go proteins ($Go{\alpha}$) and regulated neurite outgrowth triggered by $Go{\alpha}$ activation. In this study, we showed that the GTPase domain of $Go{\alpha}$ contributed to the direct interaction with Rit. We also demonstrated that $Go{\alpha}$ could lead to an increase of Rit activity suggesting that Rit play a role as a downstream effector of $Go{\alpha}$.

• Genomics & Informatics
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• v.19 no.2
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• pp.18.1-18.8
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• 2021

G protein–coupled receptors (GPCRs), including olfactory receptors, account for the largest group of genes in the human genome and occupy a very important position in signaling systems. Although olfactory receptors, which belong to the broader category of GPCRs, play an important role in monitoring the organism's surroundings, their actual three-dimensional structure has not yet been determined. Therefore, the specific details of the molecular interactions between the receptor and the ligand remain unclear. In this report, the interactions between human olfactory receptor 1A1 and its odorant molecules were simulated using computational methods, and we explored how the chemically simple odorant molecules activate the olfactory receptor.

• BMB Reports
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• v.54 no.7
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• pp.380-385
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• 2021

Proper targeting of the βPAK-interacting exchange factor (βPIX)/G protein-coupled receptor kinase-interacting target protein (GIT) complex into distinct cellular compartments is essential for its diverse functions including neurite extension and synaptogenesis. However, the mechanism for translocation of this complex is still unknown. In the present study, we reported that the conventional kinesin, called kinesin-1, can transport the βPIX/GIT complex. Additionally, βPIX bind to KIF5A, a neuronal isoform of kinesin-1 heavy chain, but not KIF1 and KIF3. Mapping analysis revealed that the tail of KIF5s and LZ domain of βPIX were the respective binding domains. Silencing KIF5A or the expression of a variety of mutant forms of KIF5A inhibited βPIX targeting the neurite tips in PC12 cells. Furthermore, truncated mutants of βPIX without LZ domain did not interact with KIF5A, and were unable to target the neurite tips in PC12 cells. These results defined kinesin-1 as a motor protein of βPIX, and may provide new insights into βPIX/GIT complex-dependent neuronal pathophysiology.

• Asian-Australasian Journal of Animal Sciences
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• v.32 no.8
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• pp.1104-1111
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• 2019

Objective: The aim of this study was to detect single nucleotide polymorphisms (SNP) of G protein-coupled receptor 54 (GPR54) gene and explore association of this candidate gene with reproductive traits in Jiaxing Black sows. Methods: Six pairs of primers of the gene were designed to amplify all exons thus sequences of which were detected by means of direct sequencing and then SNP loci were scanned. The effects of SNPs on total number of piglets born (TNB), number of piglets born alive (NBA), number of still born piglets (NSB), and litter weight at birth (LWB) of Jiaxing Black sows were analyzed. Results: Three SNP loci, including T3739C, C3878T and T6789C, were identified via comparison of sequencing and two genotypes (AB, BB) at each SNP site were observed. T3739C resulted in the change of amino acid ($Leu{\rightarrow}Pro$) in corresponding protein, and C3878T resulted in synonymous mutation ($Ile{\rightarrow}Ile$). Statistical results demonstrated that allele B was the preponderant allele at the three SNP loci and Genotype BB was the preponderant genotype. Meanwhile, Chi-Square test of these three SNPs indicated that all mutation sites fitted in Hardy-Weinberg equilibrium (p>0.05). For GPR54-T3739C locus, Jiaxing Black sows with genotype BB had 1.23 TNB and 1.28 NBA (p<0.01) that were more than those with genotype AB, respectively. Jiaxing Black sows that had the first two parities with genotype BB had additional 2.23 TNB, 2.27 NBA (p<0.01), and 1.94 LWB (p<0.05) compared to those with genotype AB, respectively. However, for other two loci, no significant difference was found between TNB, NBA, NSB, and LWB, and different genotypes of Jiaxing Black sows. Conclusion: In conclusion, the polymorphisms of GPR54-T3739C locus were significantly associated to TNB, NBA, and LWB and could be used as a potential genetic marker to improve reproductive function of Jiaxing black sows.