| 1. |
- Assalauova, Dameli, et al.
(författare)
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An advanced workflow for single-particle imaging with the limited data at an X-ray free-electron laser
- 2020
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Ingår i: IUCrJ. - 2052-2525. ; 7, s. 1102-1113
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Tidskriftsartikel (refereegranskat)abstract
- An improved analysis for single-particle imaging (SPI) experiments, using the limited data, is presented here. Results are based on a study of bacteriophage PR772 performed at the Atomic, Molecular and Optical Science instrument at the Linac Coherent Light Source as part of the SPI initiative. Existing methods were modified to cope with the shortcomings of the experimental data: inaccessibility of information from half of the detector and a small fraction of single hits. The general SPI analysis workflow was upgraded with the expectation-maximization based classification of diffraction patterns and mode decomposition on the final virus-structure determination step. The presented processing pipeline allowed us to determine the 3D structure of bacteriophage PR772 without symmetry constraints with a spatial resolution of 6.9 nm. The obtained resolution was limited by the scattering intensity during the experiment and the relatively small number of single hits.
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| 2. |
- Choi, Junhong, et al.
(författare)
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2'-O-methylation in mRNA disrupts tRNA decoding during translation elongation
- 2018
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Ingår i: Nature Structural & Molecular Biology. - : Springer Nature. - 1545-9993 .- 1545-9985. ; 25:3, s. 208-216
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Tidskriftsartikel (refereegranskat)abstract
- Chemical modifications of mRNA may regulate many aspects of mRNA processing and protein synthesis. Recently, 2 '-O-methylation of nucleotides was identified as a frequent modification in translated regions of human mRNA, showing enrichment in codons for certain amino acids. Here, using single-molecule, bulk kinetics and structural methods, we show that 2 '-O-methylation within coding regions of mRNA disrupts key steps in codon reading during cognate tRNA selection. Our results suggest that 2 '-O-methylation sterically perturbs interactions of ribosomal-monitoring bases (G530, A1492 and A1493) with cognate codon-anticodon helices, thereby inhibiting downstream GTP hydrolysis by elongation factor Tu (EF-Tu) and A-site tRNA accommodation, leading to excessive rejection of cognate aminoacylated tRNAs in initial selection and proofreading. Our current and prior findings highlight how chemical modifications of mRNA tune the dynamics of protein synthesis at different steps of translation elongation.
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| 3. |
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| 4. |
- Choi, Junhong, et al.
(författare)
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N-6-methyladenosine in mRNA disrupts tRNA selection and translation-elongation dynamics
- 2016
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Ingår i: Nature Structural & Molecular Biology. - : Springer Science and Business Media LLC. - 1545-9993 .- 1545-9985. ; 23:2, s. 110-
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Tidskriftsartikel (refereegranskat)abstract
- N-6-methylation of adenosine (forming m(6)A) is the most abundant post-transcriptional modification within the coding region of mRNA, but its role during translation remains unknown. Here, we used bulk kinetic and single-molecule methods to probe the effect of m(6)A in mRNA decoding. Although m(6)A base-pairs with uridine during decoding, as shown by X-ray crystallographic analyses of Thermus thermophilus ribosomal complexes, our measurements in an Escherichia coli translation system revealed that m(6)A modification of mRNA acts as a barrier to tRNA accommodation and translation elongation. The interaction between an m(6)A-modified codon and cognate tRNA echoes the interaction between a near-cognate codon and tRNA, because delay in tRNA accommodation depends on the position and context of m(6)A within codons and on the accuracy level of translation. Overall, our results demonstrate that chemical modification of mRNA can change translational dynamics.
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| 5. |
- Dao, E. Han, et al.
(författare)
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Structure of the 30S ribosomal decoding complex at ambient temperature
- 2018
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Ingår i: RNA. - : Cold Spring Harbor Laboratory Press (CSHL). - 1355-8382 .- 1469-9001. ; 24:12, s. 1667-1676
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Tidskriftsartikel (refereegranskat)abstract
- The ribosome translates nucleotide sequences of messenger RNA to proteins through selection of cognate transfer RNA according to the genetic code. To date, structural studies of ribosomal decoding complexes yielding high-resolution data have predominantly relied on experiments performed at cryogenic temperatures. New light sources like the X-ray free electron laser (XFEL) have enabled data collection from macromolecular crystals at ambient temperature. Here, we report an X-ray crystal structure of the Therm us thermophilus 30S ribosomal subunit decoding complex to 3.45 angstrom resolution using data obtained at ambient temperature at the Linac Coherent Light Source (LCLS). We find that this ambient-temperature structure is largely consistent with existing cryogenic-temperature crystal structures, with key residues of the decoding complex exhibiting similar conformations, including adenosine residues 1492 and 1493. Minor variations were observed, namely an alternate conformation of cytosine 1397 near the mRNA channel and the A-site. Our serial crystallography experiment illustrates the amenability of ribosomal microcrystals to routine structural studies at ambient temperature, thus overcoming a long-standing experimental limitation to structural studies of RNA and RNA-protein complexes at near-physiological temperatures.
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| 6. |
- Kurta, Ruslan P., et al.
(författare)
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Correlations in Scattered X-Ray Laser Pulses Reveal Nanoscale Structural Features of Viruses
- 2017
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Ingår i: Physical Review Letters. - : American Physical Society. - 0031-9007 .- 1079-7114. ; 119:15
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Tidskriftsartikel (refereegranskat)abstract
- We use extremely bright and ultrashort pulses from an x-ray free-electron laser (XFEL) to measure correlations in x rays scattered from individual bioparticles. This allows us to go beyond the traditional crystallography and single-particle imaging approaches for structure investigations. We employ angular correlations to recover the three-dimensional (3D) structure of nanoscale viruses from x-ray diffraction data measured at the Linac Coherent Light Source. Correlations provide us with a comprehensive structural fingerprint of a 3D virus, which we use both for model-based and ab initio structure recovery. The analyses reveal a clear indication that the structure of the viruses deviates from the expected perfect icosahedral symmetry. Our results anticipate exciting opportunities for XFEL studies of the structure and dynamics of nanoscale objects by means of angular correlations.
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| 7. |
- Li, Haoyuan, et al.
(författare)
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Diffraction data from aerosolized Coliphage PR772 virus particles imaged with the Linac Coherent Light Source
- 2020
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Ingår i: Scientific Data. - : NATURE RESEARCH. - 2052-4463. ; 7:1
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Tidskriftsartikel (refereegranskat)abstract
- Single Particle Imaging (SPI) with intense coherent X-ray pulses from X-ray free-electron lasers (XFELs) has the potential to produce molecular structures without the need for crystallization or freezing. Here we present a dataset of 285,944 diffraction patterns from aerosolized Coliphage PR772 virus particles injected into the femtosecond X-ray pulses of the Linac Coherent Light Source (LCLS). Additional exposures with background information are also deposited. The diffraction data were collected at the Atomic, Molecular and Optical Science Instrument (AMO) of the LCLS in 4 experimental beam times during a period of four years. The photon energy was either 1.2 or 1.7keV and the pulse energy was between 2 and 4 mJ in a focal spot of about 1.3 mu m x 1.7 mu m full width at half maximum (FWHM). The X-ray laser pulses captured the particles in random orientations. The data offer insight into aerosolised virus particles in the gas phase, contain information relevant to improving experimental parameters, and provide a basis for developing algorithms for image analysis and reconstruction.
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| 8. |
- Munke, Anna, et al.
(författare)
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Data Descriptor : Coherent diffraction of single Rice Dwarf virus particles using hard X-rays at the Linac Coherent Light Source
- 2016
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Ingår i: Scientific Data. - : Nature Publishing Group. - 2052-4463. ; 3
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Tidskriftsartikel (refereegranskat)abstract
- Single particle diffractive imaging data from Rice Dwarf Virus (RDV) were recorded using the Coherent X-ray Imaging (CXI) instrument at the Linac Coherent Light Source (LCLS). RDV was chosen as it is a wellcharacterized model system, useful for proof-of-principle experiments, system optimization and algorithm development. RDV, an icosahedral virus of about 70 nm in diameter, was aerosolized and injected into the approximately 0.1 mu m diameter focused hard X-ray beam at the CXI instrument of LCLS. Diffraction patterns from RDV with signal to 5.9 angstrom ngstrom were recorded. The diffraction data are available through the Coherent X-ray Imaging Data Bank (CXIDB) as a resource for algorithm development, the contents of which are described here.
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| 9. |
- Orhan, Mehmet, et al.
(författare)
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Supercritical carbon dioxide application using hydantoin acrylamide for biocidal functionalization of polyester
- 2020
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Ingår i: Journal of Supercritical Fluids. - : Elsevier. - 0896-8446 .- 1872-8162. ; 165
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Tidskriftsartikel (refereegranskat)abstract
- Biocidal functionalization in polyester fibers is a really tough challenge because of the lack of tethering groups. This study indicated supercritical carbon dioxide application using N-halamine would be an alternative solution for obtaining antibacterial function on the polyester surface. Firstly, N-(2-methyl 1-(4 methyl-2,5-dioxo-imidazolidin-4 yl)propan-2-yl)acrylamide was synthesized and applied to the polyester in supercritical carbon dioxide medium, at 120 degrees C, 30 MPa for different processing times. The addition of N-halamine on the surface significantly brought antibacterial activity against E. coli. The chlorine loadings showed that 6-h exposure time was critical to obtain sufficient antibacterial activity. This treatment caused a reasonable and tolerable loss in color and mechanical properties. But, the durability to abrasion, stability, and rechargeability of oxidative chlorine, and the durability of N-halamine on the surface were remarkably good. Conclusively, it can be available to work on polyester surfaces with resource-efficient and eco-friendly supercritical carbon dioxide technique for getting more functionalization and modification. (C) 2020 Elsevier B.V. All rights reserved.
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| 10. |
- Reddy, Hemanth K. N., et al.
(författare)
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Coherent soft X-ray diffraction imaging of coliphage PR772 at the Linac coherent light source
- 2017
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Ingår i: Scientia Danica. Series H. Humanistica 4. - : Nature Publishing Group. - 1904-5506 .- 2052-4463. ; 4
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Tidskriftsartikel (refereegranskat)abstract
- Single-particle diffraction from X-ray Free Electron Lasers offers the potential for molecular structure determination without the need for crystallization. In an effort to further develop the technique, we present a dataset of coherent soft X-ray diffraction images of Coliphage PR772 virus, collected at the Atomic Molecular Optics (AMO) beamline with pnCCD detectors in the LAMP instrument at the Linac Coherent Light Source. The diameter of PR772 ranges from 65-70 nm, which is considerably smaller than the previously reported similar to 600 nm diameter Mimivirus. This reflects continued progress in XFEL-based single-particle imaging towards the single molecular imaging regime. The data set contains significantly more single particle hits than collected in previous experiments, enabling the development of improved statistical analysis, reconstruction algorithms, and quantitative metrics to determine resolution and self-consistency.
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