6MWQ

Single particle cryoEM structure of a DARPin-aldolase platform in complex with GFP

Summary for 6MWQ

• Entry DOI: 10.2210/pdb6mwq/pdb
• EMDB information: 9277
• Descriptor: DARPin, Muscle-type aldolase chimeric fusion, Green fluorescent protein (2 entities in total)
• Functional Keywords: synthetic construct, platform, single particle cryoem, small protein display, lyase-fluorescent protein complex, lyase/fluorescent protein
• Biological source: synthetic construct; More
• Total number of polymer chains: 8
• Total formula weight: 314348.52

Authors

Weaver, S.J.,Yao, Q. (deposition date: 2018-10-30, release date: 2018-11-21, Last modification date: 2024-03-13)

Primary citation

Yao, Q.,Weaver, S.J.,Mock, J.Y.,Jensen, G.J.
Fusion of DARPin to Aldolase Enables Visualization of Small Protein by Cryo-EM.
Structure, 27:1148-, 2019

PubMed Abstract: Solving protein structures by single-particle cryoelectron microscopy (cryo-EM) has become a crucial tool in structural biology. While exciting progress is being made toward the visualization of small macromolecules, the median protein size in both eukaryotes and bacteria is still beyond the reach of cryo-EM. To overcome this problem, we implemented a platform strategy in which a small protein target was rigidly attached to a large, symmetric base via a selectable adapter. Of our seven designs, the best construct used a designed ankyrin repeat protein (DARPin) rigidly fused to tetrameric rabbit muscle aldolase through a helical linker. The DARPin retained its ability to bind its target: GFP. We solved the structure of this complex to 3.0 Å resolution overall, with 5-8 Å resolution in the GFP region. As flexibility in the DARPin position limited the overall resolution of the target, we describe strategies to rigidify this element.

PubMed: 31080120
DOI: 10.1016/j.str.2019.04.003

Experimental method

ELECTRON MICROSCOPY (3 Å)