SPR of GPCRs at Biortus
Release Time:
2023-04-06 14:11
Source:
GPCRs represent the largest family encoded by the human genome, comprising 3% with 826 members. Human GPCRs are split into four major classes: A, B, C, and F. GPCRs respond to a wide variety of stimuli, ranging from photons to small proteins. As important as the recent GPCR structures have been for drug development, it is abundantly clear that solution-based or membrane compatible biophysical and biochemical tools are needed to characterize and screen potential GPCR targeting therapeutics. Biortus’s experienced team has developed various assays to help GPCR functional assessment, one of which is Surface Plasmon Resonance (SPR).
Compared to cellular based assays, in vitro assays have much higher throughput assuming enough protein can be procured, and the protein is stable throughout the assay duration. One such assay where these two factors are critical is SPR. SPR is able to give kinetic reads and affinity measurements. The main limitation for its application to GPCRs is the upper molecular weight limit and detergent compatibility. Based on our internal tests, this assay can be applied to detergent solubilized GPCRs given the correct conditions. Nanodisc stabilization is preferred not only for GPCR purification but also for SPR as it completely removes the presence of detergent in the system. Shown here is an example of A2AR, an archetypal GPCR generated entirely at Biortus by our membrane protein team, and a small molecule antagonist, theophylline; clearly demonstrating the capability of determining binding affinity and kinetics.
Besides small molecules, GPCRs are also known to be triggered by other types of stimuli, including light, sheer stress, peptides, and more. A2AR is activated by its endogenous agonist, adenosine, and plays a prominent role in cardiac function, immune system regulation, and central nervous system signaling. APJ, a member of the class A GPCR family highly expressed in gastrulation and associated with the development of heart, cardiovascular homeostasis and cardiac contractility signaling axis, plays a critical role in the regulation of normal cardiovascular function and fluid homeostasis. Here, our Assay Team was able to determine the binding affinity of APJ and its peptide agonist, Apelin-17. The relative increase in the assay window/signal intensity compared to small molecule binding as exemplified in Figure 1 is most likely due to the larger size of the peptide and increase surface area for binding.
Biortus also provides mass photometry, nanoDSF, additional biochemical assays and cellular assays that can be applied to GPCRs. Biortus is your dedicated partner for your protein needs with protein production, assay development, fragment screening, and structure determination (MicroED, X-ray Crystallography, and CryoEM) all under one roof. For new inquiries, reach out to info@wuxibiortus.com.
Biortus now offers comprehensive portfolio of off the shelf products for the above related proteins.
|
Cata. No. |
Protein |
Species |
Sequence |
Price |
|
APJ-H41C |
APJ |
Human |
M1-R330, T177N, C325L, C326M |
|
|
A2AR-H01J |
A2AR |
Human |
S6-A316, N154A |
|
|
A2AR-H41M |
A2AR |
Human |
P2-L208, S91K, E219-A316 |
For more protein products, please reach out to order@wuxibiortus.com.
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