ProFoldin: Drug Discovery, Drug Delivery, Biochemical Assays, and Membrane Protein Sciences

In collaboration with NIH, ProFoldin has successfully developed a high-throughput assay for SARS-CoV-2 RdRp. The assay can be used to

• Detect SARS-CoV-2 RdRp activity of purified enzyme samples
• Test SARS-CoV-2 RdRp inhibitors – screening compounds and IC₅₀ measurement

COVID-19 is caused by the infection of SARS-CoV-2, or severe acute respiratory syndrome coronavirus 2, a positive-sense single-stranded RNA virus. The RNA dependent RNA polymerase (RdRp) is a key drug target for this virus. The functional SARS-CoV-2 RdRP is composed of a complex of the nsp12, nsp7, and nsp8 proteins. The SARS-CoV-2 RdRP assay is based on the detection of RNA molecules synthesized by SARS-CoV-2 RdRP in the presence of a primed RNA template, ATP, UTP, GTP, and CTP. It is a fluorescence-based assay in a 384-well or 96-well plate format. The assay procedure is simple – mixing the reagents, incubating the reaction, and reading the fluorescence. It is suitable for testing compound collections of all sizes from a few inhibitors to compound libraries. It can be employed for compound screening and IC₅₀ measurement of SARS-CoV-2 RdRp inhibitors.

Also available:
Gel-Based SARS-CoV-2 RdRp Assay Kit:
The gel-based assay can be used for visualization of RNA molecules synthesized by the RNA polymerase. It can also be used to visualize enzyme inhibition by RdRp inhibitors. The gel-based assay outcome is consistent with the high-throughput fluorescence-based assay.  Therefore, it is a useful tool for confirmation of the inhibition rates of the high-throughput screening.

ProFoldin offers high-throughput assays for validated drug targets. The assays are used for inhibition tests or IC₅₀ measurement of compounds against specific drug target proteins and enzymes. The assays are in 96-well or 384-well plate format. The validated drug targets are enzymes of various biological systems, including DNA replication, gene transcription, gene regulation, cell membrane synthesis, and other metabolism processes which are essential for growth of bacterial and cancer cells or virus replication.

ProFoldin also provides enzymes, DNA, enzyme substrates, and detection dyes separately for drug target protein characterization, assay optimization, and HTS needs.

Concentration measurement is an essential need in all laboratories from life sciences to chemistry both in academia and industry. Concentration assays for biomolecules are basic for the characterization of biological processes involving enzymatic reactions or translocation of biomolecules. ProFoldin offers high-throughput assays for various molecules and ions. The assays are used for concentration measurement of chemicals, drugs and ions in pharmaceutical compounds, environmental samples, buffers, and biochemical and chemical reactions. The assays are in 96-well format and based on fluorescence or light absorbance detection.

Assays are available for various biomolecules, including DNA, protein, ADP, UDP, GDP, histidine, cysteine, other amino acids, peptides, CoA, lipids, and phospholipids. ProFoldin also offers concentration assays for various organic and inorganic molecules, ions and drug molecules, including DTT, EDTA, detergent, detergent CMC, SDS, phosphate, polyphosphate, sulfate, chloride, Zn²⁺, Cu²⁺, Co²⁺, Ni²⁺, penicillin drugs, polypeptide drugs, Cisplatin and Oxaliplatin drugs.

ProFoldin is the leading company in protein folding technology. Since 2005, ProFoldin’s protein folding columns have been widely employed to make active proteins from inclusion bodies.

Applications:

• Fold proteins from inclusion bodies
• Fold membrane proteins from inclusion bodies
• Extract membrane proteins from cell membranes
• Membrane protein reconstitution in nanodiscs
• Evaluate protein thermal stability

Highly charged 96-well plates are available for DNA or protein binding. The biological molecule-bound plates can be used to study the interactions between the bound molecule and other molecules. They can be used for ELISA or interaction studies between nucleic acids or/and proteins.

Kits are available for removal of DNA/RNA, phosphate, or salt.

Using ProFoldin’s nanotechnology, various liposomes and nanodiscs can be produced in scales suitable for all research projects and industrial production. ProFoldin offers the following liposome products:

• Liposome purification and stability assay kits
• Ready-to-load liposomes for drug delivery
• Drug-loaded liposomes
• Liposomes loaded with dyes, ions, and biomolecules
• Nanodiscs and liposomes for membrane protein reconstitution

Detergent-Free Solubilization of Membrane Proteins

Styrene-maleic acid (SMA) copolymers are broadly used for function and structure studies of membrane proteins. The significant advantages of using SMA copolymers include (1) generating a detergent-free system and (2) forming bilayer nanodiscs with phospholipids. Application of SMA copolymers opens an avenue of membrane protein extraction from cell membranes and proteoliposomes in the absence of detergent. The extracted membrane proteins are stabilized in the nanodiscs that mimic the bilayer structure of lipids in nature.

Benefits

• Structural and functional studies on membrane proteins in a near-native environment
• Prevents protein misfolding or aggregation
• Unlike detergents, which tend to strip away most or all lipids in the direct environment of a membrane protein, SMA extracts proteins in form of a lipid/protein nanodisc
• Enables the study of protein-lipid, protein-protein, and protein-ligand interactions

ProFoldin’s SMA copolymers are optimal for nanodisc formation:

• Styrene:maleic acid ratios of 2:1 or 3:1 are the best for membrane proteins
• Molecular weights of 8 kDa and 10 kDa are ideal for nanodisc formation
• Pre-hydrolyzed and ready to be dissolved in common buffers
• No need for pH adjustment

Please see Reference List