Barrel Shape and Chromophore Rigidity Predict Fluorescent-Protein Photophysics

Document Type

Article

Publication Date

2026

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Originally published in the Journal of Chemical Information and Modeling (2026) 66 (16): 10266–10276.
https://doi.org/10.1021/acs.jcim.6c01606

PubMedCentral PMCID: PMC13508766

Abstract

The 11-stranded β-barrel of fluorescent proteins (FPs) is universally conserved, yet its quantitative geometry has not been systematically characterized. We analyzed cross-sectional barrel geometry across 908 FP crystal structures in the RCSB PDB by principal component analysis (PCA)-based axis determination and convex hull analysis of protein-atom slices at the chromophore plane; 780 structures (210–245 residues, with the chromophore-containing chain selected in FP-complex cocrystals) form the canonical analysis cohort. Barrel shape, but not size, correlates with emission wavelength: red-shifted proteins have narrower, more elliptical barrels (ρ = −0.328 for minor axis, p = 2.2 × 10–17). Fluorescence quantum yield, by contrast, is not governed by barrel size: it tracks how rigidly the barrel holds the chromophore (chromophore-to-barrel B-factor ratio, ρ = −0.49 per unique FP), together with the chromophore’s ground-state planarity (ρ = −0.42) as an independent signal of comparable strength. The planarity term is most pronounced among red fluorescent proteins, which span the widest range of ground-state twist. Principal correlations survive Benjamini–Hochberg correction and partial correlation controlling for resolution. The barrel is not a passive scaffold: it constrains chromophore rigidity and thereby shapes photophysical output. The pipeline was developed with Claude (Anthropic) via Claude Code.

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