Kavli INsD Researchers' Breakthrough Unveils Potential of Directly Combining Native Mass Spectrometry and Cryo-EM
Stephan Rauschenbach and Tim Esser, in collaboration with Kavli Institute for Nanoscience Discovery (Kavli INsD) Director Professor Carol Robinson, and Associate Professor Lindsay Baker have achieved remarkable progress in their ground-breaking research, which aims to seamlessly integrate native mass spectrometry with cryo-electron microscopy (cryo-EM).
The researchers used electrospray ion-beam deposition (ES-IBD), to bring protein complexes into the gas phase, where they can be mass selected before gentle deposition onto cryogenically cooled cryo-EM grids. Their recent efforts, highlighted in a Nature Technology Feature, and published in a BioXiV preprint, succeeded in maintaining the integrity of protein structures from solution, through the mass spectrometer to the final sample grid throughout the complex process of mass-selected, soft-landing electrospray ion beam deposition (ESIBD). The team's study presents a highly detailed cryo-EM structure of β-galactosidase, an enzyme renowned for its pivotal role in sugar metabolism, which allowed to generate an atomically resolved structure.
The success of the innovative approach sparked widespread excitement and anticipation within the scientific community. As Carol Robinson explained, "Everybody can get a good structure of β-galactosidase - but not after taking it through a mass spectrometer and landing it." Despite initial challenges and scepticism, the team's findings represent a breakthrough in applying soft-landing mass spectrometry to biological structure determination.
Determination of solution and gas-phase protein structure.
A Native electrospray ion-beam deposition workflow. Ions are transferred into the gas phase using native electrospray, mass selected, and deposited with controlled energy, density, and distribution on cold grids inside the cryo shuttle. Full and mass-selected spectra of β-galactosidase are shown. B Render of cryogenically-cooled landing stage. Protein ion beam is guided to the two grid positions using an electrostatic lens system. The stage is cooling grids and shielding them from contamination and radiation. The position of the cryo shuttle inside the cryo stage is highlighted in white.
The integration of soft-landing mass spectrometry with cryoEM holds the promise of revolutionizing sample preparation for structural biology , offering a plethora of opportunities for in-depth analyses and comprehensive insights into intricate protein structures. With an emphasis on minimizing structural disruption during the landing process, Rauschenbach, Esser, and Robinson's research signifies a critical advancement in the quest for a more refined and effective approach to studying complex protein interactions and molecular dynamics and shows folded structures of gas-phase proteins never seen before.
Their efforts align with the broader vision of the Kavli Institute for Nanoscience Discovery, which aims to foster a collaborative and interdisciplinary environment for ground-breaking scientific exploration. With this recent breakthrough, the team has illuminated new paths for future research, instilling a sense of optimism and determination among peers and enthusiasts alike. As the scientific community eagerly awaits further developments in this field, Rauschenbach, Esser, and Robinson's work stands as a testament to the power of ingenuity, perseverance, and interdisciplinary collaboration in pushing the boundaries of modern scientific exploration.
Read the full article: https://doi.org/10.1101/2023.08.17.553673
Since April 2021, Oxford University's KAVLI Institute for Nanoscience Discovery is proudly serving as a hub for research groups from seven different departments spanning both the medical and physical sciences, including Associate Professor Stephan Rauschenbach's Group from the Department of Chemistry.