Membrane proteins account for approximately 20-30% of proteins encoded by the human genome and play central roles in cell signaling, substance transport, and cell recognition. Aberrant expression and dysfunction of membrane proteins are closely associated with a variety of diseases, including cancer, neurological disorders, and metabolic diseases, making them key focuses in drug discovery.
Among them, multi-pass membrane proteins—including GPCRs, ion channels, and transporters—are especially important. They control vital cellular processes and are the targets of more than 60% of FDA-approved drugs.
1. Challenges with Multi-Pass Membrane Proteins
Full-length multi-pass membrane proteins are tricky to work with. Their complex structures and reliance on a native membrane environment make them difficult to produce, purify, and study. Some of the main challenges include:
Hard to express: Due to structural complexity, these proteins often misfold or form inclusion bodies in heterologous expression systems, leading to low yields and poor functional integrity.
Difficult to purify: A large amount of non-membrane components must be removed while maintaining protein stability. Conventional detergents frequently disrupt the structure and activity of membrane proteins.
Challenging to study structurally: Multi-pass membrane proteins are difficult to crystallize and require a membrane-like environment. Although cryo-EM has made significant progress, sample preparation remains highly challenging.
Hard to test functionally: Functional assays require a membrane-mimicking environment, as conventional aqueous systems fail to replicate the native membrane state.
To overcome bottlenecks in the expression, purification, and stabilization of full-length membrane proteins—particularly those requiring a membrane-like environment due to their multi-pass structures—researchers have developed the Nanodisc preparation platform. This system constructs a stable artificial or semi-artificial lipid bilayer microenvironment, encapsulating membrane proteins in a near-native state. Nanodiscs not only greatly enhance protein stability and activity but also provide an ideal scaffold for structural analysis, functional studies, and drug screening.
Today, Nanodiscs are available in several formats, with MSP Nanodiscs, Synthetic Nanodiscs, and PeptiNanodiscs being the major ones, each offering unique advantages for membrane protein research. To fully leverage these technologies, DIMA uses the HEK293 system to produce recombinant membrane proteins that closely mimic their natural forms, supporting faster and more reliable drug discovery.
2. Types of Nanodiscs
Nanodiscs can be classified based on the stabilizing agents used. The major formats are MSP Nanodiscs, Synthetic Nanodiscs, and PeptiNanodiscs.
2.1 MSP Nanodisc
MSP nanodiscs use membrane scaffold proteins (MSPs) as stabilizers to encapsulate artificial phospholipid components and transmembrane proteins, thereby forming a nanoscale disc. The size of the nanodisc formed is determined by the MSPs used, typically ranging from 7 to 13 nm in diameter. Commonly used MSPs in nanodisc assembly include MSP1D1, MSP1D1-DH5, and MSP1E3D1. These MSPs have been extensively studied and proven effective in generating stable and functional nanodisc proteins.
Fig 1. Assembly of MSP-Nanodisc
2.2 Synthetic Nanodisc
Synthetic nanodiscs are produced using synthetic polymers, typically styrene–maleic acid copolymer (SMA) and diisobutylene–maleic acid (DIBMA). Unlike MSP nanodiscs, the assembly of synthetic nanodiscs starts directly from intact cells. The polymers solubilize the cell membrane while native phospholipids form nanodisc structures around the membrane proteins. In this process, the polymers act simultaneously as solubilizers and stabilizers, eliminating the need for additional detergents.
Fig 2. Assembly of Synthetic Nanodisc
2.3 PeptiNanodisc
PeptiNanodisc is a technology that uses specialized peptides to wrap around the hydrophobic regions of membrane proteins (similar to peptide discs), thereby protecting them from the aqueous environment. Since neither phospholipids nor detergents are present in the final protein, membrane proteins produced using the PeptiNanodisc platform can be directly applied in cell-based assays. Moreover, in theory, peptide discs can function as a “universal scaffold,” adaptable to a wide variety of membrane proteins without being restricted by protein size. This technology enables the stable reconstitution of multiple classes of membrane proteins and is particularly suitable for presenting transmembrane targets in their native conformations.
Fig 3. Assembly of PeptiNanodisc
3. Applications of Nanodiscs
Nanodiscs are transforming how researchers work with membrane proteins in drug discovery and beyond. By keeping proteins stable and functional – whether in lipid bilayers or peptide-based scaffolds – they open the door to applications ranging from structural biology to antibody screening, vaccine design, and diagnostics. In the following subsections, we outline the major application areas, highlight where Nanodiscs add the most value, and list the assays most commonly used in each case.
3.1 Membrane Protein Structure and Function Studies
Understanding how membrane proteins fold and function requires them to remain stable in near-native conditions. Nanodiscs provide that environment, enabling high-resolution structural and biophysical studies.
Common Assays:
- Cryo-EM
- NMR, SPR, ITC, and other biophysical studies: Precise measurement of binding kinetics between membrane proteins and ligands or antibodies.
3.2 Drug Target Discovery and Validation
Accurate target presentation is essential for discovering small molecules, antibodies, and peptide drugs. Nanodiscs stabilize membrane proteins in functional forms, improving screening success and reducing false negatives.
Common Assays:
- Ligand binding assays: Screen GPCR Nanodiscs to find high-affinity lead compounds.
- Antibody screening and optimization: Flow cytometry or BLI/SPR to select antibodies with high specificity and functional activity.
3.3 Vaccine and Immunogen Design
For vaccine research, Nanodiscs help preserve conformational epitopes, making them ideal for studying and presenting conformationally sensitive antigens such as viral envelope proteins and bacterial outer membrane proteins.
Common Assays:
- Vaccine development: Display of HIV Env, SARS-CoV-2 S protein, and others on Nanodiscs to induce neutralizing antibody responses.
3.4 Detection and Diagnostics
Nanodiscs improve assay sensitivity by keeping membrane proteins functional and soluble, supporting both research assays and clinical diagnostics..
Common Assays:
- ELISA, flow cytometry, Luminex: Detect target-specific antibodies.
- Companion diagnostics: Monitor patient serum antibody responses against membrane protein targets.
3.5 Biologics Quality and Functional Assessment
Nanodiscs allow biologics to be tested against membrane proteins in their functional state, supporting therapeutic development and quality control.
Common Assays:
- ADC internalization assays
- CAR-T cell positivity rate assessment
- Thermal stability analysis: Test interactions with GPCRs or ion channels
To make it easier to see which Nanodisc types are most suitable for different applications and assays, we’ve summarized the key points in the table below:
Nanodisc Applications at a Glance
| Application | Common Assays | Recommended Nanodisc Type |
|---|---|---|
| Structure & Function | Cryo-EM, NMR, SPR, ITC, | MSP, Synthetic, PeptiNanodisc (less common) |
| Drug Discovery & Validation | Ligand binding, high-throughput screening, antibody screening (flow cytometry, BLI, SPR) | MSP, Synthetic, PeptiNanodisc (ideal for cell-based assays) |
| Vaccine & Immunogen Design | Immunogen presentation, immunogenicity studies | Synthetic, PeptiNanodisc, ⚠️ MSP may cause high background |
| Detection & Diagnostics | ELISA, flow cytometry, Luminex, companion diagnostics, cell-based assays | MSP, Synthetic, PeptiNanodiscs (ideal for cell-based assays) |
| Biologics Quality & Functional Assessment | ADC internalization, CAR-T validation, thermal stability tests | MSP, Synthetic, PeptiNanodisc (ideal for cell-based functional assays) |
4. Advantages of DIMA Nanodisc Products
The DIMA Nanodisc full-length membrane protein production platform possesses a globally leading membrane protein library, with over 500 types of full-length multi-transmembrane proteins available in stock, including GPCRs, ion channels, and other important drug targets. Technically, the platform has successfully produced full-length membrane proteins with up to 24 transmembrane domains, setting a new industry record. Product validation includes SDS-PAGE, ELISA, SPR, HPLC, etc., ensuring high purity, high solubility, and high stability, while maintaining native conformation and supporting room-temperature transport.
Additionally, the platform provides flexible, customized protein expression and production services to meet diverse research needs.
Validation Data Display
- SDS-PAGE
- ELISA
- SEC-HPLC
- SPR
- Detection of CAR-T cell positivity rate
GPRC5D PeptiNanodisc (Cat.No.FLP400011) and G4S-PE protein were used to detect the positivity rate of GPRC5D CAR-T cells.
FITC-labeled CLDN18.2 PeptiNanodisc (Cat.No.FLP420014) and G4S-PE protein were used to detect the positivity rate of CLDN18.2 CAR-T cells.
