Solution Structure and Backbone Dynamics of the Photoactive Yellow Protein^,

Abstract

The solution structure of photoactive yellow protein (PYP), a photosensory protein from Ectothiorhodospira halophila, has been determined by multidimensional NMR spectroscopy. The structure consists of an open, twisted, 6-stranded, antiparallel beta-sheet, which is flanked by four alpha-helices on both sides. The final set of 26 selected structures is well-defined for the regions spanning residues Phe6-Ala16, Asp24-Ala112, and Tyr118-Val125 and displays a root-mean-square deviation, versus the average, of 0.45 A for the backbone and 0.88 A for all heavy atoms. Comparison of the solution structure with an earlier published 1.4 A crystal structure (Borgstahl, G. E. O., Williams, D. R., and Getzoff, E. D. (1995) Biochemistry 34, 6278-6287) reveals a similarity with a root-mean-square deviation of 1.77 A for the backbone for the well-defined regions. The most distinct difference in the backbone with the crystal structure is found near the N-terminus, for residues Asp19-Leu23, which corresponds to an alpha-helix in the crystal structure and to one of the poorest defined regions in the solution structure. To characterize the dynamic behavior of PYP in solution, we undertook a 15N relaxation study and measurements of hydrogen/deuterium exchange. Determination of order parameters through the model-free Lipari-Szabo approach enabled the identification of several regions of enhanced dynamics. The comparison of atomic displacements in the backbone traces of the ensemble structures, with mobility measurements from NMR, show that the poorly defined regions feature fast internal motions in the nanosecond to picosecond time scale.

Protonation/Deprotonation Reactions Triggered by Photoactivation of Photoactive Yellow Protein from Ectothiorhodospira halophila

“…His-3 is located close to the protein surface, and in the crystal structure it appears not to be involved in the hydrogen bonding responsible for the closure of the N-terminal helical lariat (9). Furthermore, in solution the five residues at the N terminus of PYP are in a disordered state (36). Therefore, in the pG state of PYP, His-3 is already exposed to solvent and thus is not likely to contribute to the observed proton uptake at acidic pH.…”

Robustness and evolvability in the functional anatomy of a PER-ARNT-SIM (PAS) domain

“…We propose that the thermodynamic fragility of the minor hydrophobic core is functionally important by priming the stimulus-induced release of the N-terminal region. NMR measurements of PYP have revealed that the two α-helices in the N-terminal region are structurally dynamic (36), suggesting that the N-terminal region is not strongly associated with the PAS domain core of PYP. This issue is relevant to signaling by PYP, because dissociation of the N-terminal region is a key event in pB formation (30,31).…”

Influence of a Joining Helix on the BLUF Domain of the YcgF Photoreceptor from Escherichia coli

“…In addition to the light-induced structural changes seen in LOV2 domains, light-induced structural changes at the extensions of photoreceptor domains might be a general theme, as they were observed for the so-called DAS motive of A. thaliana cryptochrome 1 [58] and a N-terminal helical segment of the photoactive yellow protein. [51,59,60] Based on these data, a model for the role of the Ja-helix of YcgF during signal transduction is proposed: In the darkadapted state the C-terminal a-helix interacts with the BLUF domain and moves upon illumination relative to the domain surface, similar to the Ja-helix of LOV2. [40] The reoriented BLUF Ja-helix could thereby affect the EAL domain's activity either by controlling access to the c-di-GMP binding site or by changing its dimeric association, [31] as EAL domains can dimerize as exemplified by the N-terminal EAL domain of YkuI from B. subtilis (2BAS).…”