Membrane Proteins

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Unraveling the Secrets of Membrane Proteins: Key Targets for Drug Discovery

Membrane proteins play a crucial role in many biochemical processes, making them very attractive targets for drug discovery. However, structural analysis of membrane proteins has been usually challenging due to low protein yields and their unstable nature when removed from their native environment. In the past few years, the advent of cryo-EM and advances in sample stabilization have enabled researchers to obtain structural information for these important classes of protein from less material at a faster rate than ever before. This has expanded our understanding of membrane proteins' complex structures, and has also provided insight into their function, allowing them to be investigated in greater detail. 

The most targeted group of membrane proteins is the G-protein superfamily, followed closely by the ion channels family. However, recently, much effort has been devoted towards solute carriers as well, aiming to explore new therapeutic opportunities. Advancements in structural and analytical methods have provided valuable insights into the conformational and mechanistic dynamics of membrane proteins, as well as their drug binding modalities. This knowledge paves the way for the development of targeted drugs and therapies.

Working with membrane proteins presents specific challenges, as they need  to be extracted from their native environment and stabilized for high-resolution single-particle analysis studies. One of the main hurdles in membrane protein research is the production of sufficient quantities of soluble, stable proteins. Development of robust production methods has been a continuous area of focus, and researchers are exploring various strategies, including novel expression systems, cell-free synthesis, and lipid-based systems to improve yields.

In general, multiple constructs consisting of natural variants across different species or engineered sequences with different truncations, fusion constructs, and mutations need to be screened to identify the most suitable form.  

Several methods are then used to assess the protein stability and activity, from thermal stability to functional assays.  When assessing membrane protein stability, it is often beneficial to use a combination of  techniques to obtain a comprehensive understanding of the protein's behavior in different environments and conditions. This is usually a time-consuming process, and may take several months before success is attained.

Overcome challenges studying membrane proteins with cryo-EM high resolution data collection.

Traditionally, crystallography has been a common technique for studying proteins' three-dimensional structure, but obtaining well-diffracting crystals for membrane proteins can be challenging. Cryo-EM allows researchers to visualize membrane proteins in solution, circumventing the need for crystallization.

Since 2013, when the first membrane protein structure solved by cryo-EM was reported, the number of membrane protein structures deposited in the Protein Data Bank (PDB) has rapidly increased. Recent advances in cryo-EM are also enabling the study of smaller and more challenging targets.  One advantage of cryo-EM is that it is more forgiving of conformational variability, and multiple protein conformations can be visualized and solved from one single experiment. 

Studying membrane proteins using cryo-EM can be limited by problems caused by grid preparation procedures. Traditional plunge freezing techniques may have reproducibility issues, or may generate sub-optimal grids with respect to sample distribution and behavior. Different grid types and alternative blotting-free methods have been developed to address these limitations and NIS has extensive experience with all the tools used in grid preparation. 

More and more evidence points to the fact that membrane proteins are crucial targets  for  the development of novel  therapeutics and innovative treatments for various diseases. Through the integration of different structural biology- and biology-based approaches, scientists can combine data from various sources  to create comprehensive biological models, offering valuable insights into the structural and dynamic changes that occur at different scales. As cryo-EM and other structural techniques continue to evolve, our understanding of membrane proteins and their functions within cellular processes will undoubtedly be greatly enhanced. These advancements hold tremendous potential for unlocking new avenues in biomedical research and paving the way for groundbreaking discoveries in the field of life sciences.

3D reconstruction of AcrB protein structure determination and AcrB electron density map with cryo-EM | structural biology services | nano imaging services
3D structure reconstruction (left) with electron density maps (right) of AcrB protein, determined by high resolution cryo-EM imaging and data collection.

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