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MCM in complex with dsDNA in presence of ATP.MCM in complex with dsDNA in presence of ATP.
Structural highlights
FunctionPublication Abstract from PubMedThe cryo-electron microscopy (cryoEM) method has enabled high-resolution structure determination of numerous biomolecules and complexes. Nevertheless, cryoEM sample preparation of challenging proteins and complexes, especially those with low abundance or with preferential orientation, remains a major hurdle. We developed an affinity-grid method employing monodispersed single particle streptavidin on a lipid monolayer to enhance particle absorption on the grid surface and alleviate sample exposure to the air-water interface. Using this approach, we successfully enriched the Thermococcus kodakarensis mini-chromosome maintenance complex 3 (MCM3) on cryoEM grids through biotinylation and resolved its structure. We further utilized this affinity method to tether the biotin-tagged dsDNA to selectively enrich a stable MCM3-ATP-dsDNA complex for cryoEM structure determination. Intriguingly, both MCM3 apo and dsDNA bound structures exhibit left-handed open spiral conformations, distinct from other reported MCM structures. The large open gate is sufficient to accommodate a dsDNA which could potentially be melted. The value of mspSA affinity method was further demonstrated by mitigating the issue of preferential angular distribution of HIV-1 capsid protein hexamer and RNA polymerase II elongation complex from Saccharomyces cerevisiae. Open architecture of archaea MCM and dsDNA complexes resolved using monodispersed streptavidin affinity CryoEM.,Ma J, Yi G, Ye M, MacGregor-Chatwin C, Sheng Y, Lu Y, Li M, Li Q, Wang D, Gilbert RJC, Zhang P Nat Commun. 2024 Nov 27;15(1):10304. doi: 10.1038/s41467-024-53745-w. PMID:39604363[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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