Tracking the structural dynamics of proteins in solution using time-resolved wide-angle X-ray scattering

Cammarata, Marco; Levantino, Matteo; Schotte, Friedrich; Anfinrud, Philip; Ewald, Friederike; Choi, Jungkweon; Cupane, Antonio; Wulff, Michaël; Ihee, Hyotcherl

doi:10.1038/nmeth.1255

Cited by 251 publications

(326 citation statements)

References 31 publications

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“…Our interpretation does not deviate from the previous femtosecond Fe K-edge XANES measurements on [Fe(bpy) 3 ] 2+ by Bressler and co-workers 12 and Fe L-edge XANES measurements of a related Fe spin-crossover complex Fe[(tren(py) 3 )] 2+ by Huse and co-workers. 14 Both studies showed instrument response time-limited dynamics for the formation of the HS excited state.…”

Section: As Shown Insupporting

confidence: 53%

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Femtosecond X-ray Absorption Spectroscopy at a Hard X-ray Free Electron Laser: Application to Spin Crossover Dynamics

Lemke¹,

et al. 2013

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X-ray free electron lasers (XFELs) deliver short (<100 fs) and intense (∼10 12 photons) pulses of hard X-rays, making them excellent sources for time-resolved studies. Here we show that, despite the inherent instabilities of current (SASE based) XFELs, they can be used for measuring high-quality X-ray absorption data and we report femtosecond time-resolved Xray absorption near-edge spectroscopy (XANES) measurements of a spin-crossover system, iron(II) tris(2,2′-bipyridine) in water. The data indicate that the low-spin to high-spin transition can be modeled by single-exponential kinetics convoluted with the overall time resolution. The resulting time constant is ∼160 fs. ■ INTRODUCTIONChemical reactions involve the dynamic evolution of strongly coupled electrons and nuclei on ultrafast time scales from few to a few hundred femtoseconds. Disentangling this complex dynamics has motivated a wide range of experimental and theoretical studies to obtain a more complete picture of the elementary steps underlying these processes. 1 Despite the progresses in ultrafast optical spectroscopies that can resolve dynamics of the valence shell electrons with femtosecond time resolution and in theoretical calculations of many-body dynamics, our understanding of the early chemical transformations still remains incomplete due to the lack of direct structural measurements visualizing both electronic structure and nuclear movements in real time. Information on nuclear dynamics has been obtained with time-resolved scattering with a time resolution of 100 ps on a variety of system from solutions 2,3 to single crystals. 4−6 Recently, femtoseconds studies have been published. 7−9 When the surrounding of specific atoms is of interest, recent advances have demonstrated the utility of time-resolved X-ray absorption spectroscopy (XAS) as a reliable monitor of nuclear and electronic structure during chemical reactions on both the picosecond 10,11 and, more recently, the femtosecond time scale. 12−14 Yet, most of the synchrotron-based experiments are limited to insufficient time resolution (∼100 ps in normal operating conditions) or very low flux (∼10 photons/pulse at kilohertz rate, when operated in femtosecond laser slicing mode 15 ). The development of new X-ray facilities such as X-ray free electron lasers (XFELs) provide unprecedented capabilities in terms of both X-ray pulse durations (down to tens of femtoseconds) and single pulse intensities of more than 10 10 photons when a 1 eV bandwidth monochromator is used. In the present study we exploit XFEL radiation from the Linac Coherent Light Source (LCLS), the first hard XFEL, 16 and present an experimental study of the photoexcited spin transition dynamics in aqueous ferreous bipyridine ([Fe-(bpy) 3 ] 2+ ), which proves the feasibility of femtosecond XAS at the LCLS.Transition metal complexes exhibiting photoinduced spin crossover (SCO) transition provide a good example of the electronic dynamics coupled with nuclear motions. In a

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Section: As Shown Insupporting

confidence: 53%

“…We performed the femtosecond X-ray absorption near edge structure (XANES) measurements on a 50 mM aqueous solution of [Fe(bpy) 3 ]Cl 2 at the XPP station of the LCLS (Figure 2). The experiment used a 0.1 mm thick liquid jet oriented at an angle of 45°with respect to the direction of the incident X-ray beam.…”

Section: ■ Experimental Methodsmentioning

confidence: 99%

Femtosecond X-ray Absorption Spectroscopy at a Hard X-ray Free Electron Laser: Application to Spin Crossover Dynamics

Lemke¹,

et al. 2013

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“…3 It was also demonstrated that Mb shows a detectable difference scattering curve at the time delay of 10 ns upon the CO photolysis of carbonmonoxy Mb (MbCO), as shown in Figure 1A. This result for Mb is significant for the following reason.…”

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confidence: 80%

Protein Tertiary Structural Changes Visualized by Time-Resolved X-ray Solution Scattering

Ahn

Kim

et al. 2009

J. Phys. Chem. B

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We obtained a solution structural model of myoglobin (Mb) formed upon the CO photolysis of MbCO by analyzing time-resolved X-ray solution scattering data. An experiment-restrained rigid-body molecular dynamics simulation was used to find the best model whose theoretical difference scattering curve gives a satisfactory agreement with the experimental data at the time delay of 10 ns. The obtained solution model shows structural changes similar to crystallographic models for MbCO f Mb but also displays a noticeable difference in that the N-terminus and F helix show larger structural changes.Here, we report a transient solution structural model of myoglobin (Mb) formed 10 ns after the CO photolysis of MbCO. The model was obtained by analyzing experimental timeresolved X-ray solution scattering data with an experimentrestrained rigid-body molecular dynamics (MD) simulation. The root-mean-square deviation (rmsd) between Mb and MbCO is 0.2-0.3 Å, 1,2 underscoring the high structural sensitivity of timeresolved X-ray solution scattering.Knowledge of three-dimensional structures of proteins and their temporal progression is essential for understanding protein structure-function-dynamics relationships. Using short X-ray pulses from a synchrotron, a pump-probe time-resolved X-ray solution scattering technique was developed for tracking conformational changes of proteins in solution. 3,4 The time resolution is in principle limited by the X-ray pulse width, which is currently 100 ps and will soon be improved to better than 0.1 ps upon the delivery of X-ray free electron lasers. Time-resolved X-ray solution scattering 5-7 together with time-resolved X-ray crystallography 8 can provide direct structural information, and thus complements time-resolved optical spectroscopy in the analysis of solution-phase reaction mechanisms. Moreover, timeresolved X-ray solution scattering complements time-resolved X-ray crystallography by probing the motions of proteins in their natural environment rather than in a single crystal.Tertiary/quaternary structural transition in hemoglobin (Hb) was studied in depth with the time-resolved X-ray solution technique, and the kinetics and some preliminary structural analysis were given. 3 It was also demonstrated that Mb shows a detectable difference scattering curve at the time delay of 10 ns upon the CO photolysis of carbonmonoxy Mb (MbCO), as shown in Figure 1A. This result for Mb is significant for the following reason. Hb consists of four subunits and thus largeamplitude quaternary (rearrangements of subunits) structural transitions as well as subtle tertiary (within a subunit) structural transitions are possible between liganded and unliganded states. In contrast, Mb is a single subunit and can accompany only subtle tertiary structural changes. For example, known crystallographic models of liganded and unliganded Hb show rmsd values ranging from 4 to 9 Å through the quaternary structural change, 9 but the corresponding rmsd values between known crystallographic models of Mb and MbCO are less tha...

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“…In one notable example, DEER experiments using spin-labeled frozen samples (T = 60 K) of the photoactive yellow protein showed that the distance distribution of the labels changed on exposure of the photoreceptor to light (Ramachandran et al 2011). In the same study, timeresolved pump-probe x-ray scattering (TR-SAXS/WAXS) (Cammarata et al 2008) was used to demonstrate that the radius of gyration and particle size both increased when the protein was exposed to light (Ramachandran et al 2011). Chemical shift perturbations and classical NOE data were combined with TR-SAXS/WAXS and DEER data to produce a model of the protein's dynamic light-exposed state (Ramachandran et al 2011).…”

Section: Integration Of Nmr With Other Analytical Methodsmentioning

confidence: 99%

Protein dynamics and function from solution state NMR spectroscopy

Kovermann

Rogne

Wolf‐Watz

2016

Quart. Rev. Biophys.

143

122

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Abstract. It is well-established that dynamics are central to protein function; their importance is implicitly acknowledged in the principles of the Monod, Wyman and Changeux model of binding cooperativity, which was originally proposed in 1965. Nowadays the concept of protein dynamics is formulated in terms of the energy landscape theory, which can be used to understand protein folding and conformational changes in proteins. Because protein dynamics are so important, a key to understanding protein function at the molecular level is to design experiments that allow their quantitative analysis. Nuclear magnetic resonance (NMR) spectroscopy is uniquely suited for this purpose because major advances in theory, hardware, and experimental methods have made it possible to characterize protein dynamics at an unprecedented level of detail. Unique features of NMR include the ability to quantify dynamics (i) under equilibrium conditions without external perturbations, (ii) using many probes simultaneously, and (iii) over large time intervals. Here we review NMR techniques for quantifying protein dynamics on fast (ps-ns), slow (μs-ms), and very slow (s-min) time scales. These techniques are discussed with reference to some major discoveries in protein science that have been made possible by NMR spectroscopy.

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Tracking the structural dynamics of proteins in solution using time-resolved wide-angle X-ray scattering

Cited by 251 publications

References 31 publications

Femtosecond X-ray Absorption Spectroscopy at a Hard X-ray Free Electron Laser: Application to Spin Crossover Dynamics

Femtosecond X-ray Absorption Spectroscopy at a Hard X-ray Free Electron Laser: Application to Spin Crossover Dynamics

Protein Tertiary Structural Changes Visualized by Time-Resolved X-ray Solution Scattering

Protein dynamics and function from solution state NMR spectroscopy

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