Minireview: Structural insights into early folding events using continuous‐flow time‐resolved small‐angle X‐ray scattering

Kathuria, Sagar V.; Guo, Liang; Graceffa, Rita; Barrea, Raúl A.; Nobrega, R. Paul; Matthews, C. Robert; Irving, Thomas C.; Bilsel, Osman

doi:10.1002/bip.21628

Cited by 50 publications

(43 citation statements)

References 66 publications

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“…For these studies, the protein was denatured using 4.5 M [GdnHCl] to populate a conformational state with a radius of gyration consistent with a random coil (Kathuria et al, 2011). A tenfold dilution of this solution with buffer in the continuous-flow mixer gives rise to conditions favoring the native state.…”

Section: Resultsmentioning

confidence: 99%

“…One important application is the study of protein and RNA folding, processes that involve a large dynamic range in both length and time scales, with many important processes involved in hydrophobic collapse occurring on the microsecond to millisecond time scale (Kathuria et al, 2011;Sosnick & Barrick, 2011;Svergun & Koch, 2003;Thirumalai et al, 2001;Woodson, 2010). Similarly, ligand and RNA/DNA binding (Wee et al, 2012), assembly of lipid bilayer structures and nano-particle-based drug delivery systems (Johnson & Prud'homme, 2003), vesicle formation (Weiss et al, 2005;Guida, 2010), protein association (Doyle et al, 2004) and conformational dynamics (Chattopadhyay et al, 2002;Werner et al, 2006;Srajer & Royer, 2008) also typically occur on a sub-millisecond timescale.…”

Section: Introductionmentioning

confidence: 99%

“…Various kinds of continuous-flow mixers, however, have the potential to go much faster (Kathuria et al, 2011). Among these approaches, turbulent mixing methods (Roder et al, 2004) have the advantage of being able to take advantage of all of the delivered flux in the X-ray beam for the best signalto-noise ratio (S/N).…”

Section: Introductionmentioning

confidence: 99%

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Sub-millisecond time-resolved SAXS using a continuous-flow mixer and X-ray microbeam

et al. 2013

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Small-angle X-ray scattering (SAXS) is a well established technique to probe the nanoscale structure and interactions in soft matter. It allows one to study the structure of native particles in near physiological environments and to analyze structural changes in response to variations in external conditions. The combination of microfluidics and SAXS provides a powerful tool to investigate dynamic processes on a molecular level with sub-millisecond time resolution. Reaction kinetics in the sub-millisecond time range has been achieved using continuous-flow mixers manufactured using micromachining techniques. The time resolution of these devices has previously been limited, in part, by the X-ray beam sizes delivered by typical SAXS beamlines. These limitations can be overcome using optics to focus X-rays to the micrometer size range providing that beam divergence and photon flux suitable for performing SAXS experiments can be maintained. Such micro-SAXS in combination with microfluidic devices would be an attractive probe for time-resolved studies. Here, the development of a high-duty-cycle scanning microsecond-timeresolution SAXS capability, built around the Kirkpatrick-Baez mirror-based microbeam system at the Biophysics Collaborative Access Team (BioCAT) beamline 18ID at the Advanced Photon Source, Argonne National Laboratory, is reported. A detailed description of the microbeam small-angle-scattering instrument, the turbulent flow mixer, as well as the data acquisition and control and analysis software is provided. Results are presented where this apparatus was used to study the folding of cytochrome c. Future prospects for this technique are discussed.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Sub-millisecond time-resolved SAXS using a continuous-flow mixer and X-ray microbeam

et al. 2013

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“…5has been taken from the work ofBrennich et al (2011) who studied the dynamics of intermediate filament assembly using a continuous micromixer. SAXS data were collected at different positions in the jet, observing the lateral aggregation development and salt concentration dependence of the assembly Kathuria et al (2011). recently presented a review on the study of early events in protein folding using continuous mixers and SAXS.…”

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confidence: 99%

Time-resolved structure investigation with small angle X-ray scattering using scanning techniques

Amenitsch

Marmiroli

2011

Rend. Fis. Acc. Lincei

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Due to the availability of high-brilliance X-rays at synchrotron radiation facilities, small angle X-ray scattering (SAXS) measurements have become feasible for new sample environments, like microfluidics or aerosol systems. The combination of continuous flow methods and SAXS with spatial scanning enables in situ measurements of time-resolved structures at the nanoscale important for reactions kinetics and material synthesis and processing. In this review we present the latest achievements in terms of scanning SAXS along continuous flow mixing devices reducing the limit of time resolution at the microsecond range. This led to the determination of early stages of biological reactions like protein folding, and to gain insight into chemical reaction kinetics like nanoparticles formation. We also describe the coupling of scanning SAXS with aerosol generators, which is the only way to study the mechanisms for the self-assembly and aggregation of nanomaterials via the aerosol route. We finally give a brief outlook, as the potentiality of these techniques has not been fully exploited, leaving the possibility for further improvements in time resolution and sample consumption, for example with the new X-ray sources like Free Electron Lasers.

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“…The microfluidic environment offers precise control of the magnitude and geometry of the flow fields [11][12][13][14][15][16] and may be combined with online synchrotron Small Angle X-ray Scattering (SAXS) to follow changes in the lyotropic phases under flow. Despite the challenges of coupling X-ray scattering with microdevices, due to material compatiblity and scattering background, 17,18 successful experiments have been carried out, including the resolution of ms-conformational changes in proteins and RNA, [19][20][21] alignment of anisotropic particles and liquid crystals, 22,23 phase transitions in block copolymer solutions 24 The structural changes in the spatially varying flows, analogous to the processing and manufacturing routes in a tubular reactor, can be correlated with the vis-coelastic properties of the complex fluid to elucidate the interplay of flow history and rheological properties of polymer solutions 19,20,25 and surfactant mixtures. [26][27][28][29][30][31] This microfluidic approach is thus complementary to well-established rheo-SAXS and stopped-flow techniques, 4,32 providing unprecedented spatio-temporal insight into the flow response of complex fluids.…”

Section: Introductionmentioning

confidence: 99%

Microfluidic processing of concentrated surfactant mixtures: online SAXS, microscopy and rheology

et al. 2016

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We investigate the effect of microfluidic flow on the microstructure and dynamics of a model surfactant mixture, combining synchrotron Small Angle X-ray Scattering (SAXS), microscopy and rheology. A system comprising a single-chain cationic surfactant, hexadecyl trimethyl ammonium chloride (C 16 TAC), a short-chain alcohol (1-pentanol) and water was selected for the study due to its flow responsiveness and industrial relevance. Model flow fields, including sequential contraction-expansion (extensional) and rotational flows, were investigated and the fluid response in terms of the lamellar d-spacing, orientation and birefringence was monitored in situ, as well as the recovery processes after cessation of flow. Extensional flows are found to result in considerable d-spacing increase (from approx 59Å to 65Å). However, under continuous flow, swelling decreases with increasing flow velocity, eventually approaching the equilibrium values at velocities 2 cms −1 . Through individual constrictions we observe the alignment of lamellae along the flow velocity, accompanied by increasing birefringence, followed by an orientation flip whereby lamellae exit perpendicularly to the flow direction. The resulting microstructures are mapped quantitatively onto the flow field in 2D with 200 µm spatial resolution. Rotational flows alone do not result in appreciable changes in lamellar spacing and flow type and magnitude evidently impact the fluid microstructure under flow, as well as upon relaxation. The findings are correlated with rheological properties measured ex-situ to provide a mechanistic understanding of the effect of flow imposed by tubular processing units in the phase behavior and performance of a model surfactant system with ubiquitous applications in personal care and coatings industries.

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Minireview: Structural insights into early folding events using continuous‐flow time‐resolved small‐angle X‐ray scattering

Cited by 50 publications

References 66 publications

Sub-millisecond time-resolved SAXS using a continuous-flow mixer and X-ray microbeam

Sub-millisecond time-resolved SAXS using a continuous-flow mixer and X-ray microbeam

Time-resolved structure investigation with small angle X-ray scattering using scanning techniques

Microfluidic processing of concentrated surfactant mixtures: online SAXS, microscopy and rheology

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