Tue 20 Apr 11:00: LMB Seminar: Molecular views into cellular functions by in-cell cryo-electron tomography

LMB Seminar: Molecular views into cellular functions by in-cell cryo-electron tomography

Most structural biology focuses on the structure and function of individual macromolecular complexes, but falls short of revealing how they come together to give rise to cellular functions. As a consequence, structural and cell biology have traditionally been separate disciplines and employed techniques that were well defined within the realm of either one or the other. Here, cryo-electron tomography (cryo-ET) provides a unique opportunity for obtaining structural information across a wide range of spatial scales – from whole cells to individual macromolecules. We develop and employ advanced sample preparation techniques for in-cell cryo-electron tomography, including cryo-focused ion beam thinning guided by 3D correlative fluorescence microscopy. Preparations of site-specific ‘electron-transparent windows’ in cellular model systems enable assignment of molecular structures directly from three-dimensional stills of intact cells and reveal their molecular sociology. Using the genome-reduced human pathogen Mycoplasma pneumoniae as a model system, we further developed the synergistic application of whole-cell crosslinking mass spectrometry, cellular cryo-ET and integrative modelling, and determine in-cell structures of transient, actively transcribing RNA polymerases coupled to a translating ribosomes. Recent computational breakthroughs now allow resolving these molecular machines to residue-level and reveal small molecule antibiotics bound to their active cite within the intact pathogen. These methodologies unlock an enormous potential for novel discovery enabled by label-free structural cell biology.

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