Chromophore charge-state switching through copper-dependent homodimerisation of an engineered green fluorescent protein.

ABSTRACT: Here, we have linked one of the most common protein-protein interaction events, homodimerisation, to an essential trace metal, copper, through engineering green fluorescent protein. Mutation of H148 to cysteine promotes the neutral phenolic chromophore in the monomer that excites predominantly at ∼400 nm. Homodimerisation via a copper-dependent disulphide bridge switches the chromophore to the charged phenolate that excites at ∼490 nm. The result is a ∼30 fold increase in the fluorescence emission ratio. Homodimerisation kinetics are further improved by optimising the sfGFP homodimer interface, generating the variant termed GFP-diS2. Structures of the monomeric and dimeric GFP-diS2 suggest that charge switching is through peptide bond flipping and the formation of a buried organised

SUBMITTER: Ahmed RD 

PROVIDER: S-EPMC12547689 | biostudies-literature | 2025 Nov

REPOSITORIES: biostudies-literature