Protein solubilization is a pivotal step in the protein production process, influencing the quality and functionality of custom proteins. The primary goal of protein solubilization is to convert insoluble or partially soluble proteins into a soluble form. At Eurofins CALIXAR, we recognize the critical role this step plays in unlocking the full potential of custom protein. The process involves optimizing conditions to enhance the protein solubility, ensuring the production of fully native and functional proteins, for both membrane and soluble proteins — a cornerstone for various successful applications, such as antibody discovery, vaccine formulation, drug development, bioassay development, and structural biology studies.
Eurofins CALIXAR employs cutting-edge techniques in protein solubilization, offering unmatched solutions for both membrane and soluble proteins. In our custom protein services, our innovative solubilization solutions stand as cores in our commitment to delivering high-quality, fully native, and functional proteins. We provide two techniques, including innovative detergent and synthetic nanodiscs.
Unlocking Protein Potential: A Step-by-Step Guide to Solubilization
- Method Selection: The journey begins with the strategic choice between traditional detergents and innovative synthetic nanodiscs. The selection depends on the unique characteristics of the protein of interest.
- Sample Preparation: Ensure meticulous sample preparation, considering the source of the protein and the intended application. This lays the groundwork for a successful solubilization process.
- Detergent Screening: If utilizing detergents, embark on a screening process to identify the most suitable detergent. This step ensures compatibility and efficient solubilization of membrane proteins.
- Synthetic Nanodisc Formation: Opt for synthetic nanodiscs as an alternative, initiating the formation process using carefully selected polymers like SMA, AASTY, or Ultrasolute™ Amphipol. These nanodiscs seamlessly blend solubilization and stabilization.
- Incubation: Allow the protein sample to incubate with the chosen solubilization agent, providing adequate time for the interaction between the agent and the hydrophobic regions of the protein.
- Optimization of Conditions: Fine-tune conditions such as solubilization time, temperature, and buffer composition to optimize the process, enhancing protein solubility and stability.
- Isolation of Solubilized Proteins: Employ separation techniques like centrifugation or filtration to isolate the solubilized proteins, capturing the desired soluble form while leaving insoluble components behind.
Unveiling the essence of protein solubilization at Eurofins CALIXAR
1. Innovative detergent
Our detergents, a classic approach, represent an advanced approach to solubilizing proteins. At Eurofins CALIXAR, we undertake an accurate screening process to identify the most suitable detergents tailored to your specific protein. This screening encompasses parameters such as detergent concentration, solubilization time, and buffer composition, ensuring optimal conditions for solubilization, superior stability and functionality. Customized to meet the unique demands of each project, these detergents enhance the solubility of both membrane and soluble proteins.
2. Synthetic nanodiscs
Synthetic nanodiscs are nanoscale structures that are designed to solubilize and stabilize membrane proteins in a lipid bilayer environment. Membrane proteins are a class of proteins that are embedded in the lipid bilayer of cell membranes, and they play crucial roles in various biological processes. However, studying and manipulating membrane proteins can be challenging due to their hydrophobic nature and the need for a lipid environment. Synthetic nanodiscs offer an alternative approach for solubilizing membrane proteins while maintaining a more native-like lipid environment. They provide a controlled and customizable environment for studying membrane proteins, allowing researchers to investigate their structure, function, and interactions with other molecules.
The use of synthetic nanodiscs in protein solubilization has gained popularity in structural biology and biophysical studies, as they provide a more native-like environment compared to traditional detergent-based methods. The mobile, near-identical environment they provide mimics native cell membrane, enabling a more accurate representation of the natural conditions in which membrane protein functions. These nanodiscs, increasingly replacing detergents, seamlessly combine solubilization and stabilization steps.
Synthetic nanodiscs, formed by polymers like SMA, Amphipol and DIBMA, have revolutionized membrane protein solubilization.
What are SMA polymers?
SMA (styrene-maleic acid) polymers are a class of amphipathic copolymers that have gained attention in the field of membrane protein solubilization, particularly in the context of synthetic nanodiscs. SMA polymers can form nanodisc-like structures called styrene-maleic acid lipid particles (SMALPs).
The process of nanodiscs formation using of SMA copolymers to extract membrane proteins from lipid bilayers:
- Selection of SMA Copolymer: Choose an appropriate SMA copolymer based on the specific requirements of the experiment. The copolymer typically consists of alternating styrene and maleic acid monomer units.
- Preparation of Lipid Nanodisc Assembly: Combine the SMA copolymer with the lipid bilayer containing the membrane protein of interest.
- Solubilization of Lipid Bilayer: SMA copolymers have an amphiphilic nature, with hydrophobic styrene units and hydrophilic maleic acid units. When added to a lipid bilayer, SMA disrupts the lipid packing, solubilizing the membrane and incorporating the lipid into the nanodisc.
- Extraction of Membrane Proteins: The amphipathic nature of SMA helps in extracting membrane proteins from the lipid bilayer, incorporating them into the developing nanodisc structure.
- Formation of Nanodiscs: The interaction of SMA copolymers with lipids and membrane proteins leads to the self-assembly of nanodisc structures. These nanodiscs typically consist of a lipid bilayer encircled by the SMA copolymer.
- Removal of Excess Copolymer and Lipids: Purify the nanodiscs by removing excess SMA copolymer and lipids. This can be achieved through techniques like size-exclusion chromatography.
- Characterization: Characterize the resulting nanodiscs to ensure the presence of the desired membrane protein and evaluate the size and stability of the nanodiscs.
The advantages of SMA polymers for nanodisc formation include their ability to work in physiological conditions, their relatively mild extraction process, and their compatibility with a wide range of membrane proteins:
- Native-like Lipid Environment: SMALPs provide a more native-like lipid environment for membrane proteins, preserving their structural and functional integrity.
- Mild Extraction Conditions:The use of SMA involves a mild extraction process, preventing denaturation or loss of function in membrane proteins.
- Physiological Relevance: SMA is compatible with physiological conditions, allowing researchers to study membrane proteins in a biologically relevant environment.
- Broad Applicability: SMA has demonstrated effectiveness with a diverse range of membrane proteins, making it versatile for various research fields.
- Ease of Use: The one-step process of SMALP formation simplifies the solubilization process, saving time and resources.
- Avoidance of Detergent Interference: SMA eliminates concerns related to detergent interference, providing a detergent-free environment for downstream applications.
- Stabilization of Membrane Proteins: SMA aids in stabilizing membrane proteins during solubilization, ensuring the production of high-quality protein samples.
This approach has been employed in various structural and functional studies of membrane proteins, facilitating research in areas such as structural biology, biophysics, and drug discovery. The use of SMA polymers and SMALPs has expanded the toolkit available for researchers studying membrane proteins and has provided a valuable alternative to conventional solubilization methods.
Unlock potential of native and functional proteins
The use of our innovative detergents or synthetic nanodiscs like SMA polymers offers a powerful and versatile method for membrane protein solubilization. This approach not only provides a more native-like environment but also simplifies the extraction process, making it an attractive choice for various applications in structural biology, biophysics, and drug discovery.
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