6c9k

Single-Particle reconstruction of DARP14 - A designed protein scaffold displaying ~17kDa DARPin proteins

Loading... Status: waiting for Mol* initialization Structural highlights 6c9k is a 24 chain structure with sequence from Pseae and Synthetic construct sequences. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance. Related: 6c9i Gene: PA1966 (PSEAE) Resources: FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT Publication Abstract from PubMed Current single-particle cryo-electron microscopy (cryo-EM) techniques can produce images of large protein assemblies and macromolecular complexes at atomic level detail without the need for crystal growth. However, proteins of smaller size, typical of those found throughout the cell, are not presently amenable to detailed structural elucidation by cryo-EM. Here we use protein design to create a modular, symmetrical scaffolding system to make protein molecules of typical size suitable for cryo-EM. Using a rigid continuous alpha helical linker, we connect a small 17-kDa protein (DARPin) to a protein subunit that was designed to self-assemble into a cage with cubic symmetry. We show that the resulting construct is amenable to structural analysis by single-particle cryo-EM, allowing us to identify and solve the structure of the attached small protein at near-atomic detail, ranging from 3.5- to 5-A resolution. The result demonstrates that proteins considerably smaller than the theoretical limit of 50 kDa for cryo-EM can be visualized clearly when arrayed in a rigid fashion on a symmetric designed protein scaffold. Furthermore, because the amino acid sequence of a DARPin can be chosen to confer tight binding to various other protein or nucleic acid molecules, the system provides a future route for imaging diverse macromolecules, potentially broadening the application of cryo-EM to proteins of typical size in the cell. Near-atomic cryo-EM imaging of a small protein displayed on a designed scaffolding system.,Liu Y, Gonen S, Gonen T, Yeates TO Proc Natl Acad Sci U S A. 2018 Mar 5. pii: 1718825115. doi:, 10.1073/pnas.1718825115. PMID:29507202 From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.

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