Genomics and Applied Biology 2024, Vol.15, No.1, 27-38 http://bioscipublisher.com/index.php/gab 31 tool for researchers to explore the structure and function of proteins in different biological systems. 2.2 Wide applicability Cryo EM technology plays an important role in the field of protein structure analysis due to its unique advantages, especially its wide range of applications, highlighting the value of this technology. Shoemaker and Ando (2018) delved into the significant advantage of cooling without sample crystallization, which overcomes the main obstacles in traditional X-ray crystallography and enables the structural analysis of proteins or macromolecular complexes that are difficult to crystallize. This technology is particularly suitable for studying large molecular complexes, such as viruses, ribosomes, and membrane protein complexes, which play crucial roles in biological processes and can be observed and analyzed under conditions close to their physiological states. By rapidly freezing samples at extremely low temperatures, cryo electron microscopy can lock molecules in their almost pristine state, preserving their true dynamic structure and morphology. Baker et al. (2017) discussed how to use electron freeze chromatography (cryoET) of frozen hydration samples to determine the structure of macromolecular complexes in their native environment, thereby avoiding possible sample damage or structural changes during crystal formation. This ability to maintain the original ecological state of biomolecules, combined with high-resolution imaging of large molecules and complex assembly structures, provides a powerful tool for a deeper understanding of biological mechanisms. The flexibility and diversity of cryo electron microscopy technology have further expanded its applicability, enabling it to capture the dynamic processes of molecular transitions between different states and reveal how molecules perform biological functions by changing their shape. Trabuco et al. (2009) developed the Molecular Dynamics Flexible Fit (MDFF) method to combine high-resolution atomic structures with cryo EM images, representing the atomic model of the conformational states captured by cryo EM. This method has been successfully applied to ribosomes, a ribonucleoprotein complex responsible for protein synthesis. Combined with other biophysical and biochemical technologies, cryo electron microscopy can provide comprehensive information on the structure, dynamics, and function of biomolecules, thereby promoting a comprehensive understanding of biological processes from atomic level to cellular and even tissue level. These advantages make cryo electron microscopy a powerful tool for understanding the mysteries of life sciences, and its wide applicability provides unlimited possibilities for biomedical research and drug development. 2.3 Ability to analyze proteins with different conformations Freezing electron microscopy technology is particularly adept at handling sample heterogeneity, able to identify different conformational states from tens of thousands of protein molecule images. This single particle analysis (SPA) technique is crucial for understanding how proteins participate in biological processes in different conformational states. Freezing electron microscopy does not require protein crystallization and can be directly analyzed in its natural state, which is particularly important for proteins that are difficult to crystallize and have multiple conformations. Ki et al. (2021) introduced the nanoparticle assisted cryo EM sampling (NACS) method to access the conformational distribution of protein molecules. By measuring the distance between two gold nanoparticles as a structural parameter, various protein conformations can be captured even for small or disordered proteins that are typically inaccessible through cryo EM. Jonić Reviewed with Vénien Buryan the methods and latest developments of cryo EM for structural analysis of protein and biomolecular complexes. These complexes are either too large or too heterogeneous to be studied through traditional X-ray crystallography or nuclear magnetic resonance (NMR), and these latest developments provide exciting opportunities for determining the three-dimensional structure of macromolecular complexes. Nakane et al. (2020) obtained human membrane proteins using a novel electronic source, energy filter, and camera β The 1.7 Å resolution cryo EM reconstruction of 3 GABA A receptors provides a true atomic resolution view,
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