Tryptophan fluorescence monitors structural changes accompanying signalling state formation in the photocycle of photoactive yellow protein

Publisher Website
Google Scholar
Abstract
Photoactive yellow protein, a small, water-soluble blue-light absorbing photoreceptor protein from Ectothiorhodospira (Halorhodospira) halophila has a structure with two hydrophobic cores, of which the main one houses its light-sensitive chromophore (p-coumaric acid), separated by a central β-sheet. This photoreceptor protein contains a single tryptophan residue (W119) that is situated at the interface between the central β-sheet and its N-terminal cap. The fluorescence properties of W119 in the dark state pG (λmax = 328 nm; Φfl = 0.01; nearly pH-independent) are typical for a buried tryptophan in a hydrophobic environment with significant quenching by nearby amino acid residues. Signalling state formation leads to pH-dependent fluorescence changes: At pH values in vivo, these changes lead to an exposure of the part of the central β-sheet harbouring W119. At slightly acidic pH the polarity of the environment of W119 is hardly affected by the formation of the signalling state but the quenching of its fluorescence emission, possibly by nearby amino acids, is reduced. On the other hand, its accessibility for quenching by small molecules in the solution is enhanced at acidic and alkaline pH in the signalling state (pB) compared to the dark state (pG). This latter observation points towards a more flexible structure of the N-terminal cap, having a looser interaction with the central β-sheet in pB.