G protein-coupled receptor (GPCR) phosphorylation is a pivotal regulatory mechanism in cellular signaling, influencing receptor activity, internalization, and interactions with downstream effectors. The detection of GPCR phosphorylation level provides critical insights into physiological and pathological processes, enabling diagnostic development, drug discovery, and therapeutic optimization. Phosphorylation dynamics of GPCRs is essential for evaluating drug efficacy, resistance, and therapeutic targeting.
Amerigo Scientific offers fast, robust, and reliable assay kits using immunoassay for the quantitative assessment of GPCR phosphorylation. These phosphorylation assays involve immunoprecipitation of the affinity-labeled receptor using magnetic beads followed by detection with phosphosite-specific antibodies as phosphorylation biosensors to detect receptor activation, which can be performed entirely in multi-well plates without western blot analysis.
Product Range
Adrenoceptors
Adrenoceptors are classified into α and β subtypes, belonging to the GPCR family. Upon activation, G protein-coupled receptor kinases (GRKs) phosphorylate the intracellular domains of the receptor. The phosphorylated receptor binds to β-arrestin, leading to receptor desensitization and internalization. This process plays a critical role in regulating cardiovascular function, metabolism, neurotransmission, and other physiological processes, ensuring appropriate cellular responses to epinephrine and norepinephrine. The assessment of adrenoceptor phosphorylation level can help elucidate the efficacy of therapeutics and guide novel drug development.
Bombesin Receptors
Bombesin receptors including BB1, BB2, and BB3 subtypes, are members of the GPCR family. They are activated by the endogenous ligands gastrin-releasing peptide (GRP) and neuromedin B (NMB). The BB phosphorylation regulates cell proliferation, secretion, neuroregulation, gastrointestinal function, central nervous system activity, and tumorigenesis. Recent studies have shown that BB3 plays an important role in metabolic homeostasis and obesity, and thus may be a potential target for the treatment of obesity.
Cannabinoid Receptors
Cannabinoid receptors are a class of G protein-coupled seven-transmembrane-spanning receptor proteins, including CB1 and CB2 subtypes. They are phosphorylated by GRKs upon activation by endogenous or exogenous cannabinoids. Their phosphorylation is involved in the regulation of neurotransmission, immune responses, pain perception, and energy metabolism to maintain homeostasis. CB1 receptors are preferentially expressed in the brain and central nervous system, while CB2 receptors are exclusively expressed in immune cells and peripheral nerve terminals. Antagonists of CB1 have an antiparkinsonian effect and antagonism of CB1 may be a treatment for obesity, multiple sclerosis, Alzheimer's disease, etc. Although its biological function is still unclear, CB2 receptor appear to be responsible for the anti-inflammatory and possibly other therapeutic effects of cannabis.
Chemerin Chemokine-Like Receptors
Chemokine-like receptor 1 (CMKLR1) also known as ChemR23 undergoes phosphorylation, leading to desensitization and internalization, thereby regulating chemerin-mediated inflammatory signaling. This process modulates immune cell chemotaxis, inflammatory responses, and adipocyte differentiation. Phosphorylation analysis of CMKLR1 facilitates research on Inflammatory diseases, pains, metabolic syndrome, and cardiovascular disorders, aiding the development of anti-inflammatory and metabolic-regulating drugs.
Complement Component Receptors
Complement system is a group of serum molecules that mediate inflammation and opsonization of antigens and microorganisms in addition to controlling lysis of pathogens or cells sensitized with antibody. C3a, C5a and C5adesArg are a small peptide fragments that are produced by the complement cascade. These complement components are potent chemoattractants and secretagogues that act on immune and non-immune cells. Their physiological effects are mediated by a family of three seven transmembrane receptors (7TMRs) including C3AR, C5AR1 and C5AR2.
Dopamine Receptors
Dopamine receptors are G-protein-coupled seven-transmembrane-spanning receptor proteins and include D1, D2, D3, D4, and D5 receptors. D1 receptors which stimulate the synthesis of cyclic AMP, and D2 receptors which inhibit cyclic AMP synthesis were first identified. The dopamine D3 and D4 receptors are homologous to the D2 receptor, and the dopamine D5 receptor is homologous to the D1 receptor. Dopamine receptors are phosphorylated by GRKs and PKC, which regulates motor control, emotion, endocrine function, and cognition. Activation of the D1 receptor is involved in regulation of food intake. The D2 receptor is a target for the treatment of disorders associated with dopaminergic dysfunctions such as Parkinson’s disease and schizophrenia. D3 receptor antagonists may be effective as a treatment for anxiety.
Formylpeptide Receptors
Formylpeptide receptors (FPRs) are seven transmembrane domain GPCRs expressed on neutrophils or monocytes. They are phosphorylated upon bacterial formylpeptide fMet-Leu-Phe (fMLP) activation, modulating chemotaxis, inflammatory responses, and immune defense. The human genome contains 3 genes coding for a subfamily of three FPRs. The primary function of FPR1 is recognition of bacterial mitochondrial formylpeptides. FPR2 can interact with a variety of ligands with different structures including lipid mediators such as lipoxin A4. Phosphorylation studies of FPRs can contribute to understanding the mechanisms of infection and inflammatory diseases, as well as the development of anti-infection and anti-inflammatory drugs.
Neuropeptide Y Receptors
Neuropeptide Y receptors are a subgroup of GPCRs that binds to neuropeptide Y (NPY), peptide YY (PYY), and pancreatic polypeptide, which are divided into Y1, Y2, Y4 and Y5 subtypes. Neuropeptide Y is a 36-amino acid neurotransmitter present in the brain and autonomic nervous system. It has been shown to inhibit presynaptic excitatory transmission through suppression of calcium channels and activation of potassium channels, suggesting potential utility in the treatment of epilepsy. NPY is also a central appetite stimulant that promotes initiation of feeding and delays its termination. The Y1, Y2, and Y5 receptors preferentially bind NPY and PYY, whereas the Y4 receptor preferentially binds pancreatic polypeptide. Y2 may be implicated in Huntington's disease, morbid obesity, and obesity, and is the biomarker of peripheral artery disease and temporal lobe epilepsy.
Opioid Receptors
Opioid receptors are a class of GPCRs expressed peripherally and centrally. They are abundant in regions of the brain that bind morphine as well as in areas that are unrelated and related to pain. Opioid receptors are divided into μ-opioid receptor (mu-opioid receptor, MOP), δ-opioid receptor (delta-opioid receptor, DOP), κ-opioid receptor (kappa-opioid receptor, KOP), and opioid-like 1 (ORL1) receptors. Natural ligands for these receptors are the opiate peptide neurotransmitters derived from proopiomelanocortin, proenkephalin, prodynorphin, or pronociceptin/orphanin FQ.
Agonists of the MOP are used in the treatment of abdominal pain in irritable bowel syndrome and antagonists of this receptor together with levodopa may be effective in symptomatic treatment of Parkinson's disease without eliciting dyskinesias. The DOP agonists may be effective in the treatment of the symptoms of Parkinson's disease, and KOP has showed neuroprotective effects against ischemic neuronal injury and agonists may be effective treatment of stroke. Agonists to the mu-, delta- and kappa-opioids receptors may be effective as a treatment for chronic pain disorders. Phosphorylation assays for opioid receptors can enhance understanding of pain management, opioid tolerance, and addiction mechanisms, as well as related drug discovery and development.
Somatostatin Receptors
Somatostatin is a tetradecapeptide that inhibits the release of growth hormone (GH) and thyroid stimulating hormone (TSH) from the pituitary as well as release of insulin and gastrin from the pancreas. Endogenous somatostatin has an extremely short half-life, and somatostatin analogues are thus developed to improve on the pharmacokinetic profile of the endogenous substance. Somatostatin acts by binding to G-protein-coupled somatostatin receptors, of which five subtypes are known to exist (SSTR1-SSTR5). Five subtypes are expressed to varying degrees on somatotroph adenoma cells, and thus are a target for the treatment of acromegaly. Drugs acting on somatostatin receptors have potent antisecretory and tumor volume-reducing effects. Phosphorylation studies of somatostatin receptors can advance research on endocrine diseases, tumors, and neurological diseases, and contributes to the development of somatostatin analogues.
Sphingosine 1-Phosphate Receptors
Sphingosine-1-phosphate (S1P), also known as lysosphingolipid (LPL), is a signaling sphingolipid that is characterized by sphingosine. It is released from ceramides via ceramidase-mediated catalysis. S1P is an extracellular ligand for the S1P receptor family which is a group of GPCRs important for lipid signaling. There are eight S1P receptors, including S1P1 (edge1); LPA1 (edg2); S1P3 (edg3); LPA2 (edg4); S1P2 (edg5); S1P4 (edg6); LPA3 (edg7); and S1P5 (edg8). These high-affinity GPCRs are widely expressed throughout the body and are involved in several physiologic activities including cell migration and proliferation, angiogenesis, and neurogenesis. Inhibition of S1P receptors is critical for immunomodulation and targeting of S1P receptors could be effective in controlling the autoimmune diseases.
Tachykinin Receptors
The tachykinins are a family of polypeptides or neurokinins including substance P (SP), neurokinin A (NKA), neuromedin L, substance K, neuropeptide K (NPK), neuropeptide gamma (NPγ), and neurokinin B (NKB). Their biological effects are mediated via three tachykinin receptor subtypes, neurokinin-1 (NK1), NK2, and NK3, which are members of the superfamily of G-protein-coupled, seven transmembrane-spanning receptors. SP is the preferred ligand for the NK1 receptor, while NKA and NKB are the preferred ligands for NK2 and NK3 receptors, respectively. NK1 and NK3 antagonists are under development for the treatment of anxiety and depression.
Thyrotropin-Releasing Hormone (TRH) Receptor
Thyrotropin-releasing hormone (TRH) receptors are seven transmembrane domain GPCRs, which are divided into three subtypes, TRHR1, TRHR2, and TRHR3. TRH receptors are activated by binding to the endogenous tripeptide TRH. TRH is synthesized in the hypothalamus and released into the hypothalamic-pituitary portal circulation to act on the pituitary as a neurotransmitter/neuromodulator. TRH receptor phosphorylation regulates TSH release, impacting thyroid function, metabolic rate, growth, and energy balance.
Vasopressin Receptors
Vasopressin is a peptide hormone also known as antidiuretic hormone (ADH) that is synthesized in the hypothalamus and released from the posterior pituitary. This hormone has antidiuretic and vasopressor actions. Vasopressin activity is mediated via V1 (consisting of V1A and V1B subtypes) and V2 receptors, and antagonists to either may be effective in the treatment of hypertension. In addition, circulating arginine vasopressin (AVP) is a potent vasoconstrictor, and high levels of AVP may contribute to systemic vasoconstriction and hyponatremia, making this an attractive target for the treatment of both acute and chronic heart failure. V1A receptors are located on blood vessels, where they exert vasoconstrictor effects, and in the myocardium, where they act as a myocardial stimulant. V2 receptors mediate the effects of AVP on water retention. V1B receptors have only indirect circulatory effects and thus do not constitute an important target.
VIP/PACAP Receptors
Pituitary adenylate cyclase activating polypeptide (PACAP) is a 27 - or 38-amino acid neuropeptide belonging to the vasoactive intestinal peptide (VIP)/glucagon/secretin family, which shows particularly high homology to VIP. PACAP regulates a variety of physiological functions in the peripheral and central nervous systems through the GPCRs including PAC1 receptor (PAC1-R), VPAC1 receptor (VPAC1-R), and VPAC2 receptor (VPAC2-R), which have similar affinity for PACAP and VIP. Novel therapeutics targeting PACAP is in development for the treatment of migraine.
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