Detergent-Free GPCR-Fc Fusion Binds with S-Layer-Protein A/G on Conducting Surfaces and Uses Thereof


This technology enables the drug discovery for therapeutics targeting G-protein-coupled receptors, and can be applied in the fields of biotechnology and pharmaceuticals.

Problem Addressed

Humans express nearly 1000 different G-protein-coupled receptors (GPCRs), which are plasma membrane-embedded signaling proteins that coordinate cell-cell interactions, protein trafficking, cellular migration, and metabolism, among other functions. When the activity of a GPCR becomes dysregulated, tumor initiation, progression, invasion, and metastasis can result. Consequently, GPCRs are critical therapeutic targets. It is estimated that one-half to one-third of all marketed drugs act through binding a GPCR. Nevertheless, there remain many GPCRs for which ligands and inhibitors have not yet been identified.

GPCRs are transmembrane proteins, which makes their isolation for use in ligand-binding assays challenging. Transmembrane proteins are difficult to solubilize, extract, and purify. When detergent is used to solubilize the GPCRs, the detergent can impair protein structure, stability, and function. The present technology is a cell-free, detergent-free platform and method that facilitates ligand-binding assays for water-soluble variants of GPCRs.


This invention is a self-assembling two-dimensional (2D) crystal lattice formed by surface layer (S-layer) proteins bound to modified, water-soluble GPCR variants. Here, fusion domains have been attached to the C-terminus of S-layer peptides. To avoid the use of detergents to confer water-solubility to the GPCR, the GPCR protein of interest is altered such that all hydrophobic amino acid residues found in the 7 α-helical transmembrane domains of the protein are replaced with hydrophilic residues. The inventors have created a system that replaces distinct hydrophobic residues with hydrophilic residues of a similar size to maintain GPCR structure and function. For example, the hydrophobic phenylalanine residue contains a benzene ring, which can be replaced by tyrosine, which contains a phenol group. Phenol groups are benzene rings which have been modified with a hydroxyl group, thus conferring hydrophilicity without significantly altering the size of the residue. Therefore, this amino acid replacement confers increased water-solubility without impacting protein structure.

These water-soluble variant GPCRs are fused to a binding moiety at the C-terminus. The C-terminal binding moiety of the GPCR has binding affinity for the S-layer protein fusion domain, which allows the S-layer fusion protein and the modified GPCR to associate. The S-layer fusion proteins are able to self-assemble into a crystal lattice containing water-soluble GPCR variants. This crystallized complex can assemble onto bioelectronic interfaces to serve as a platform for high-throughput GPCR ligand-binding assays to uncover novel molecules that may act as GPCR agonists/antagonists.


  • Cell-free method for testing ligand-binding for GPCRs
  • Amino acid replacement method maintains the integrity of the GPCR protein without the need for detergent
  • Method can be applied to a wide variety of GPCRs with unknown ligands to advance drug discovery

Intellectual Property

IP Type: Published US Patent Application

IP Title: S-layer protein 2D lattice coupled detergent-free GPCR bioelectronic interfaces, devices, and methods for the use thereof

IP Number: US 2019-0353654


IP Type: Published PCT Application

IP Title: S-layer protein 2D lattice coupled detergent-free GPCR bioelectronic interfaces, devices, and methods for the use thereof

IP Number: WO 2019-046699