scholarly journals Structure of the 30S ribosomal decoding complex at ambient temperature

2018 ◽  
Author(s):  
E. Han Dao ◽  
Frédéric Poitevin ◽  
Raymond G. Sierra ◽  
Cornelius Gati ◽  
Yashas Rao ◽  
...  
Keyword(s):  
Ambient Temperature ◽  
Messenger Rna ◽  
Thermus Thermophilus ◽  
Ribosomal Subunit ◽  
Temperature Structure ◽  
Structural Studies ◽  
High Resolution Data ◽  
X Ray ◽  
Experimental Limitation ◽  
Using Data

ABSTRACTThe 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 lightsources 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 Thermus thermophilus 30S ribosomal subunit decoding complex to 3.45 Å 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.

Structural studies of the 30S subunit of ribosomes Thermus thermophilus by small-angle neutron and X-ray scattering

2000 ◽  
Vol 33 (3) ◽  
pp. 515-518
Author(s):  
L. Fan ◽  
D. I. Svergun ◽  
V. V. Volkov ◽  
V. L. Aksenov ◽  
C. Ch. Algalarov ◽  
...  
Keyword(s):  
Small Angle ◽  
Thermus Thermophilus ◽  
Structural Studies ◽  
X Ray ◽  
X Ray Scattering ◽  
30S Subunit ◽  
Ray Scattering

scholarly journals Yes, one can obtain better quality structures from routine X-ray data collection

IUCrJ ◽  
2016 ◽  
Vol 3 (1) ◽  
pp. 61-70 ◽  
Author(s):  
W. Fabiola Sanjuan-Szklarz ◽  
Anna A. Hoser ◽  
Matthias Gutmann ◽  
Anders Østergaard Madsen ◽  
Krzysztof Woźniak
Keyword(s):  
Structural Parameters ◽  
Critical Discussion ◽  
Thermal Parameters ◽  
Geometrical Parameters ◽  
Vibrational Entropy ◽  
Structural Investigations ◽  
High Resolution Data ◽  
X Ray ◽  
Atom Model ◽  
Using Data

Single-crystal X-ray diffraction structural results for benzidine dihydrochloride, hydrated and protonated N,N,N,N-peri(dimethylamino)naphthalene chloride, triptycene, dichlorodimethyltriptycene and decamethylferrocene have been analysed. A critical discussion of the dependence of structural and thermal parameters on resolution for these compounds is presented. Results of refinements against X-ray data, cut off to different resolutions from the high-resolution data files, are compared to structural models derived from neutron diffraction experiments. The Independent Atom Model (IAM) and the Transferable Aspherical Atom Model (TAAM) are tested. The average differences between the X-ray and neutron structural parameters (with the exception of valence angles defined by H atoms) decrease with the increasing 2θmax angle. The scale of differences between X-ray and neutron geometrical parameters can be significantly reduced when data are collected to the higher, than commonly used, 2θmax diffraction angles (for Mo Kα 2θmax > 65°). The final structural and thermal parameters obtained for the studied compounds using TAAM refinement are in better agreement with the neutron values than the IAM results for all resolutions and all compounds. By using TAAM, it is still possible to obtain accurate results even from low-resolution X-ray data. This is particularly important as TAAM is easy to apply and can routinely be used to improve the quality of structural investigations [Dominiak (2015). LSDB from UBDB. University of Buffalo, USA]. We can recommend that, in order to obtain more adequate (more accurate and precise) structural and displacement parameters during the IAM model refinement, data should be collected up to the larger diffraction angles, at least, for Mo Kα radiation to 2θmax = 65° (sin θmax/λ < 0.75 Å−1). The TAAM approach is a very good option to obtain more adequate results even using data collected to the lower 2θmax angles. Also the results of translation–libration–screw (TLS) analysis and vibrational entropy values are more reliable for 2θmax > 65°.

scholarly journals Sarecycline interferes with tRNA accommodation and tethers mRNA to the 70S ribosome

2020 ◽  
Vol 117 (34) ◽  
pp. 20530-20537
Author(s):  
Zahra Batool ◽  
Ivan B. Lomakin ◽  
Yury S. Polikanov ◽  
Christopher G. Bunick
Keyword(s):  
Messenger Rna ◽  
Acne Vulgaris ◽  
Thermus Thermophilus ◽  
Ribosomal Subunit ◽  
Direct Interaction ◽  
Bacterial Ribosome ◽  
70S Ribosome ◽  
A Site ◽  
Aminoacyl Transfer

Sarecycline is a new narrow-spectrum tetracycline-class antibiotic approved for the treatment of acne vulgaris. Tetracyclines share a common four-ring naphthacene core and inhibit protein synthesis by interacting with the 70S bacterial ribosome. Sarecycline is distinguished chemically from other tetracyclines because it has a 7-[[methoxy(methyl)amino]methyl] group attached at the C7 position of ring D. To investigate the functional role of this C7 moiety, we determined the X-ray crystal structure of sarecycline bound to theThermus thermophilus70S ribosome. Our 2.8-Å resolution structure revealed that sarecycline binds at the canonical tetracycline binding site located in the decoding center of the small ribosomal subunit. Importantly, unlike other tetracyclines, the unique C7 extension of sarecycline extends into the messenger RNA (mRNA) channel to form a direct interaction with the A-site codon to possibly interfere with mRNA movement through the channel and/or disrupt A-site codon–anticodon interaction. Based on our biochemical studies, sarecycline appears to be a more potent initiation inhibitor compared to other tetracyclines, possibly due to drug interactions with the mRNA, thereby blocking accommodation of the first aminoacyl transfer RNA (tRNA) into the A site. Overall, our structural and biochemical findings rationalize the role of the unique C7 moiety of sarecycline in antibiotic action.

Difference Fourier Analysis of Glucose Embedded and Frozen Hydrated Purple Membrane

1982 ◽  
Vol 40 ◽  
pp. 74-75
Author(s):  
Jules S. Jaffe ◽  
Robert M. Glaeser
Keyword(s):  
Purple Membrane ◽  
Data Set ◽  
High Resolution Data ◽  
X Ray ◽  
X Ray Crystallography ◽  
Fourier Techniques ◽  
Versus Protein ◽  
The Difference ◽  
Difference Fourier ◽  
Ideal Method

Although difference Fourier techniques are standard in X-ray crystallography it has only been very recently that electron crystallographers have been able to take advantage of this method. We have combined a high resolution data set for frozen glucose embedded Purple Membrane (PM) with a data set collected from PM prepared in the frozen hydrated state in order to visualize any differences in structure due to the different methods of preparation. The increased contrast between protein-ice versus protein-glucose may prove to be an advantage of the frozen hydrated technique for visualizing those parts of bacteriorhodopsin that are embedded in glucose. In addition, surface groups of the protein may be disordered in glucose and ordered in the frozen state. The sensitivity of the difference Fourier technique to small changes in structure provides an ideal method for testing this hypothesis.

Ribosome Structural Organization

1974 ◽  
Vol 32 ◽  
pp. 332-333
Author(s):  
James A. Lake
Keyword(s):  
Ribosomal Proteins ◽  
Structural Organization ◽  
Three Dimensional ◽  
Small Subunit ◽  
Structural Studies ◽  
X Ray Diffraction ◽  
Specific Antibodies ◽  
X Ray ◽  
E Coli ◽  
Complete Sequences

The understanding of ribosome structure has advanced considerably in the last several years. Biochemists have characterized the constituent proteins and rRNA's of ribosomes. Complete sequences have been determined for some ribosomal proteins and specific antibodies have been prepared against all E. coli small subunit proteins. In addition, a number of naturally occuring systems of three dimensional ribosome crystals which are suitable for structural studies have been observed in eukaryotes. Although the crystals are, in general, too small for X-ray diffraction, their size is ideal for electron microscopy.

Quantitative x-ray microanalysis and thickness determination using ζ factor

1996 ◽  
Vol 54 ◽  
pp. 580-581
Author(s):  
M. Watanabe ◽  
Z. Horita ◽  
M. Nemoto
Keyword(s):  
Diffusion Couple ◽  
Extrapolation Method ◽  
Thin Specimen ◽  
Beam Position ◽  
X Ray ◽  
Thickness Determination ◽  
Experimental Limitation ◽  
X Ray Absorption

X-ray absorption in quantitative x-ray microanalysis of thin specimens may be corrected without knowledge of thickness when the extrapolation method or the differential x-ray absorption (DXA) method is used. However, there is an experimental limitation involved in each method. In this study, a method is proposed to overcome such a limitation. The method is developed by introducing the ζ factor and by combining the extrapolation method and DXA method. The method using the ζ factor, which is called the ζ-DXA method in this study, is applied to diffusion-couple experiments in the Ni-Al system.For a thin specimen where incident electrons are fully transparent, the characteristic x-ray intensity generated from a beam position, I, may be represented as I = (NρW/A)Qωaist.

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