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Publication Abstract from PubMed

Recent technical advances have revolutionized the field of cryo-electron microscopy (cryo-EM). However, most monomeric proteins remain too small (<100 kDa) for cryo-EM analysis. To overcome this limitation, we explored a strategy whereby a monomeric target protein is genetically fused to a homo-oligomeric scaffold protein and the junction optimized to allow the target to adopt the scaffold symmetry, thereby generating a chimeric particle suitable for cryo-EM. To demonstrate the concept, we fused maltose-binding protein (MBP), a 40 kDa monomer, to glutamine synthetase, a dodecamer formed by two hexameric rings. Chimeric constructs with different junction lengths were screened by biophysical analysis and negative-stain EM. The optimal construct yielded a cryo-EM reconstruction that revealed the MBP structure at sub-nanometre resolution. These findings illustrate the feasibility of using homo-oligomeric scaffolds to enable cryo-EM analysis of monomeric proteins, paving the way for applying this strategy to challenging structures resistant to crystallographic and NMR analysis.

Fusion to a homo-oligomeric scaffold allows cryo-EM analysis of a small protein.,Coscia F, Estrozi LF, Hans F, Malet H, Noirclerc-Savoye M, Schoehn G, Petosa C Sci Rep. 2016 Aug 3;6:30909. doi: 10.1038/srep30909. PMID:27485862^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.