Abstract
BackgroundProteins in their majority act rarely as single entities. Multisubunit macromolecular complexes are the actors in most of the cellular processes. These nanomachines are hold together by weak protein-protein interactions and undergo functionally important conformational changes. TFIID is such a multiprotein complex acting in eukaryotic transcription initiation. This complex is first to be recruited to the promoter of the genes and triggers the formation of the transcription preinitiation complex involving RNA polymerase II which leads to gene transcription. The exact role of TFIID in this process is not yet understood.MethodsLast generation electron microscopes, improved data collection and new image analysis tools made it possible to obtain structural information of biological molecules at atomic resolution. Cryo-electron microscopy of vitrified samples visualizes proteins in a fully hydrated, close to native state. Molecular images are recorded at liquid nitrogen temperature in low electron dose conditions to reduce radiation damage. Digital image analysis of these noisy images aims at improving the signal-to-noise ratio, at separating distinct molecular views and at reconstructing a three-dimensional model of the biological particle.ResultsUsing these methods we showed the early events of an activated transcription initiation process. We explored the interaction of the TFIID coactivator with the yeast Rap1 activator, the transcription factor TFIIA and the promoter DNA. We demonstrated that TFIID serves as an assembly platform for transient protein-protein interactions, which are essential for transcription initiation.ConclusionsRecent developments in electron microscopy have provided new insights into the structural organization and the dynamic reorganization of large macromolecular complexes. Examples of near-atomic resolutions exist but the molecular flexibility of macromolecular complexes remains the limiting factor in most case. Electron microscopy has the potential to provide both structural and dynamic information of biological assemblies in order to understand the molecular mechanisms of their functions.
Highlights
IntroductionMultisubunit macromolecular complexes are the actors in most of the cellular processes
Proteins in their majority act rarely as single entities
Cryo-EM of frozen hydrated molecular complexes Imaging of single particles by electron microscopy and numerical analysis of image datasets have proven invaluable tools to describe the structural organization of large macromolecular assemblies
Summary
Multisubunit macromolecular complexes are the actors in most of the cellular processes These nanomachines are hold together by weak protein-protein interactions and undergo functionally important conformational changes. In most of the cases, conformational changes that range from atomic to molecular scale are instrumental to explain the function of these complexes These dynamic properties, associated with the size of the particles ranging between 10 and 40 nm substantiates the name of nanomachines often attributed to these complexes. Few other examples of drugs targeting so clearly the intrinsic mechanical properties of a complex were described so far This is related to the poor structural information available to date on complexes since most of the atomic structures deposited in the protein data bank are single polypeptides
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