Discovery of a buried charge-network GFP-fold family spanning prokaryotes and eukaryotes
TL;DR
Findings expand the known sequence and chemical space of the GFP barrel, although the biochemical function of the network remains unknown, and identify the same two-cation/two-anion/tyrosine network in 238 members of DUF2490, an unrelated outer-membrane beta-barrel family consistent with convergent evolution of the buried network.
Abstract
The GFP fold has long been viewed as a specialized fluorescent scaffold found principally in marine animals. A structure-first search reveals a rare buried charge-network GFP-fold family spanning bacteria, fungi, and corals. Its members retain the eleven-stranded barrel and conserved chromophore-forming machinery but replace the canonical central tyrosine with methionine or other hydrophobic residues. Ancestral reconstruction supports a non-tyrosine state early in the lineage. Instead of a conventional chromophore, the barrel encloses an invariant five-residue network of two arginines, two glutamates, and Tyr168, whereas the chromophore-equivalent residue varies. The network is more conserved than the protein surface, occupies the most strongly constrained evolutionary sites, and remains formed across independent structure predictions and explicit-solvent molecular-dynamics trajectories. Fold-independent searches identify the same two-cation/two-anion/tyrosine network in 238 members of DUF2490, an unrelated outer-membrane beta-barrel family, consistent with convergent evolution of the buried network. These findings expand the known sequence and chemical space of the GFP barrel, although the biochemical function of the network remains unknown.