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 β-sheet, which is flanked by four α-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 Å for the backbone and 0.88 Å for all heavy atoms. Comparison of the solution structure with an earlier published 1.4 Å 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 Å 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 α-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.