High-speed raster-scanning synchrotron serial microcrystallography with a high-precision piezo-scanner

Gao, Yuan; Xu, Weihe; Shi, Wuxian; Soares, Alexei S.; Jakoncic, Jean; Myers, Stuart; Martins, Bruno; Skinner, John M.; Li, Qun; Bernstein, H. J.; McSweeney, Seán; Nazaretski, Evgeny; Fuchs, Martin R.

doi:10.1107/s1600577518010354

Cited by 19 publications

(26 citation statements)

References 54 publications

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“…Steps of crystal identification and rotation data collection could be combined as originally proposed and tested (Gati et al, 2014). This mode of data collection reduces unnecessary radiation damage but with a compromise of having a lot of empty frames that have to be identified and rejected prior to standard diffraction data analysis (Gao et al, 2018).…”

Section: Microdiffraction Data Collectionmentioning

confidence: 99%

“…Synchrotron beamlines optimized for microdiffraction have become available for routine user access (Perrakis et al, 1999;Flot et al, 2010;Smith et al, 2012;Fuchs et al, 2016;Yamamoto et al, 2017). Microcrystal synchrotron crystallography is now becoming increasingly attractive at microdiffraction beamlines (Ji et al, 2010;Zeldin et al, 2013a;Gati et al, 2014;Stellato et al, 2014;Botha et al, 2015;Coquelle et al, 2015;Nogly et al, 2015;Beyerlein et al, 2017;Diederichs & Wang, 2017;Martin-Garcia et al, 2017;Meents et al, 2017;Gao et al, 2018). Using crystals of 10-20 mm in size, synchrotron SAD phasing has been demonstrated with higher Z elements including iodine (Z = 53) and selenium (Z = 34) as well as for low-Z native-SAD (Melnikov et al, 2017;Huang et al, 2018).…”

Section: Introductionmentioning

confidence: 99%

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Synchrotron microcrystal native-SAD phasing at a low energy

Guo

Zhu

Fuchs

et al. 2019

IUCrJ

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De novo structural evaluation of native biomolecules from single-wavelength anomalous diffraction (SAD) is a challenge because of the weakness of the anomalous scattering. The anomalous scattering from relevant native elements – primarily sulfur in proteins and phosphorus in nucleic acids – increases as the X-ray energy decreases toward their K-edge transitions. Thus, measurements at a lowered X-ray energy are promising for making native SAD routine and robust. For microcrystals with sizes less than 10 µm, native-SAD phasing at synchrotron microdiffraction beamlines is even more challenging because of difficulties in sample manipulation, diffraction data collection and data analysis. Native-SAD analysis from microcrystals by using X-ray free-electron lasers has been demonstrated but has required use of thousands of thousands of microcrystals to achieve the necessary accuracy. Here it is shown that by exploitation of anomalous microdiffraction signals obtained at 5 keV, by the use of polyimide wellmounts, and by an iterative crystal and frame-rejection method, microcrystal native-SAD phasing is possible from as few as about 1 200 crystals. Our results show the utility of low-energy native-SAD phasing with microcrystals at synchrotron microdiffraction beamlines.

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Section: Microdiffraction Data Collectionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Synchrotron microcrystal native-SAD phasing at a low energy

Guo

Zhu

Fuchs

et al. 2019

IUCrJ

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“…The crystals for multicrystal data collection ranged in sizes from 10 × 30 × 10 µm to 30 × 50 × 15 µm (Figure 3b). Diffraction data were collected via the raster scan data collection protocol described previously [16]. The area was scanned five times with starting oscillation angles of 10°, 40°, 70°, 100°, and 130°, with the loop face perpendicular to the X-ray beam at the starting angle of 70°.…”

Section: Resultsmentioning

confidence: 99%

“…In contrast, serial data collection at synchrotrons allows for rotational data collection and multiple exposures of a single crystal due to the reduced X-ray intensity [15]. Advances in instrumentation at the FMX beamline allow for the integration of raster scanning data collection of multiple microcrystals mounted across the surface of a loop or mesh [16].…”

Section: Introductionmentioning

confidence: 99%

Getting the Most Out of Your Crystals: Data Collection at the New High-Flux, Microfocus MX Beamlines at NSLS-II

et al. 2019

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Advances in synchrotron technology are changing the landscape of macromolecular crystallography. The two recently opened beamlines at NSLS-II—AMX and FMX—deliver high-flux microfocus beams that open new possibilities for crystallographic data collection. They are equipped with state-of-the-art experimental stations and automation to allow data collection on previously intractable crystals. Optimized data collection strategies allow users to tailor crystal positioning to optimally distribute the X-ray dose over its volume. Vector data collection allows the user to define a linear trajectory along a well diffracting volume of the crystal and perform rotational data collection while moving along the vector. This is particularly well suited to long, thin crystals. We describe vector data collection of three proteins—Akt1, PI3Kα, and CDP-Chase—to demonstrate its application and utility. For smaller crystals, we describe two methods for multicrystal data collection in a single loop, either manually selecting multiple centers (using H108A-PHM as an example), or “raster-collect”, a more automated approach for a larger number of crystals (using CDP-Chase as an example).

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“…Compared with XFELs which produce only one diffraction pattern for every microcrystal, synchrotron microdiffraction beamlines are optimized for collection of a small wedge of rotation data from each microcrystal, thus greatly improving data quality from microcrystals (Smith et al, 2012;Fuchs et al, 2016;Yamamoto et al, 2017). With the implementation of new data collection methods, microcrystal data acquisition at synchrotrons is now routine and maturing (Gati et al, 2014;Coquelle et al, 2015;Zander et al, 2015;Diederichs & Wang, 2017;Owen et al, 2017;Sanishvili & Fischetti, 2017;Gao et al, 2018;Huang et al, 2018;Basu et al, 2019;Cianci et al, 2019;Dauter, 2019;Guo et al, 2019).…”

Section: Introductionmentioning

confidence: 99%

PyMDA: microcrystal data assembly using Python

et al. 2020

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The recent developments at microdiffraction X-ray beamlines are making microcrystals of macromolecules appealing subjects for routine structural analysis. Microcrystal diffraction data collected at synchrotron microdiffraction beamlines may be radiation damaged with incomplete data per microcrystal and with unit-cell variations. A multi-stage data assembly method has previously been designed for microcrystal synchrotron crystallography. Here the strategy has been implemented as a Python program for microcrystal data assembly (PyMDA). PyMDA optimizes microcrystal data quality including weak anomalous signals through iterative crystal and frame rejections. Beyond microcrystals, PyMDA may be applicable for assembling data sets from larger crystals for improved data quality.

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High-speed raster-scanning synchrotron serial microcrystallography with a high-precision piezo-scanner

Abstract: The design and first measurements with a high-speed piezo-positioner-based goniometer for raster-scanning serial macromolecular micro-focus crystallography at synchrotron storage rings are presented.

Cited by 19 publications

References 54 publications

Synchrotron microcrystal native-SAD phasing at a low energy

Synchrotron microcrystal native-SAD phasing at a low energy

Getting the Most Out of Your Crystals: Data Collection at the New High-Flux, Microfocus MX Beamlines at NSLS-II

PyMDA: microcrystal data assembly using Python

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