7UGT

Crystal structure of hyperfolder fluorescent protein FOLD6

Summary for 7UGT

• Entry DOI: 10.2210/pdb7ugt/pdb
• Descriptor: FOLD6 (2 entities in total)
• Functional Keywords: hyperfolder, fluorescent protein, superfolder, gfp, hfyfp
• Biological source: Aequorea victoria
• Total number of polymer chains: 1
• Total formula weight: 25844.06

Authors

Campbell, B.C.,Liu, C.F.,Petsko, G.A. (deposition date: 2022-03-25, release date: 2022-10-26, Last modification date: 2024-11-06)

Primary citation

Campbell, B.C.,Paez-Segala, M.G.,Looger, L.L.,Petsko, G.A.,Liu, C.F.
Chemically stable fluorescent proteins for advanced microscopy.
Nat.Methods, 19:1612-1621, 2022

PubMed Abstract: We report the rational engineering of a remarkably stable yellow fluorescent protein (YFP), 'hyperfolder YFP' (hfYFP), that withstands chaotropic conditions that denature most biological structures within seconds, including superfolder green fluorescent protein (GFP). hfYFP contains no cysteines, is chloride insensitive and tolerates aldehyde and osmium tetroxide fixation better than common fluorescent proteins, enabling its use in expansion and electron microscopies. We solved crystal structures of hfYFP (to 1.7-Å resolution), a monomeric variant, monomeric hyperfolder YFP (1.6 Å) and an mGreenLantern mutant (1.2 Å), and then rationally engineered highly stable 405-nm-excitable GFPs, large Stokes shift (LSS) monomeric GFP (LSSmGFP) and LSSA12 from these structures. Lastly, we directly exploited the chemical stability of hfYFP and LSSmGFP by devising a fluorescence-assisted protein purification strategy enabling all steps of denaturing affinity chromatography to be visualized using ultraviolet or blue light. hfYFP and LSSmGFP represent a new generation of robustly stable fluorescent proteins developed for advanced biotechnological applications.

PubMed: 36344833
DOI: 10.1038/s41592-022-01660-7

Experimental method

X-RAY DIFFRACTION (1.21 Å)