Membrane protein

Membrane proteins pose significant challenges in the field of structural biology, requiring substantial investments in terms of time and financial resources. However, the ability to quickly determine the most suitable expression system and purification conditions using minimal material can greatly reduce the initial research time and financial burden, thereby accelerating the early stages of a project.

At Biortus, we developed our small-scale workflows designed to facilitate membrane protein expression and purification screening. Here, we offer support in rapidly cloning and screening constructs simultaneously on a small scale (3-5ml of cell culture per condition) using various expression systems such as E.coli, yeast, insect or mammalian cell cultures. To expedite the process, we employ commonly used purification tags (e.g., His, FLAG, Twin-Strep, GST) and reporter proteins (e.g., eGFP, RFP, YFP, mStayGold) to quickly identify the most suitable expression and purification conditions. This includes optimizing factors such as incubation time and temperature, solvent composition, detergents, pH, salts, and other additives.

We assess the success of the screening process by utilizing techniques such as fluorescence size exclusion chromatography (FSEC), SDS-PAGE gels, Western Blot. To address the challenge of extremely low expression levels of membrane proteins, we have the capability to utilize anti-eGFP nanobody resin, which offers high specificity in capturing eGFP-tagged membrane proteins (with a dissociation constant, kd, of ~6 pM). We have successfully purified over 300 unique membrane proteins such as GPCR, ion channel, transporters by using this approach. We have a variety of membrane proteins available for purchase online. For more information, please contact us.

Our membrane protein production platform enables initial purification on a small scale, and also provide clients with the opportunity to screen a broader range of conditions. This includes, but is not limited to, detergents, amphipols, membrane scaffold proteins (MSPs), SMALPS, lipids, pH, and salts.

Maintaining the functional integrity of purified membrane proteins is crucial for downstream applications. At Biortus, we offer a range of biochemical assays, such as ATPase assays for ABC transporters, as well as biophysical assays including thermal shift assays, surface plasmon resonance (SPR), and TRIC (temperature related intensity change) assays for the purified membrane proteins. These assays help assess the functionality and stability of the proteins, ensuring their suitability for further studies and applications.

At Biortus, we also specialize in Cryo-Electron Microscopy (Cryo-EM) studies of membrane proteins. Our expertise extends to the determination of various types of membrane proteins, including inactive state G-protein coupled receptors (GPCRs), active state GPCRs, ion channels, solute carrier proteins (SLCs), and ATP-binding cassette (ABC) transporters. By combining our expertise in Cryo-EM with other biochemical and biophysical assays, we can provide comprehensive insights into the structure and function of membrane proteins. This information is crucial for understanding their roles in cellular processes and for the development of targeted therapeutics.