scholarly journals Small-Angle X-ray Scattering Models of APOBEC3B Catalytic Domain in a Complex with a Single-Stranded DNA Inhibitor

Viruses ◽  
2021 ◽  
Vol 13 (2) ◽  
pp. 290
Author(s):  
Fareeda M. Barzak ◽  
Timothy M. Ryan ◽  
Maksim V. Kvach ◽  
Harikrishnan M. Kurup ◽  
Hideki Aihara ◽  
...  
Keyword(s):  
Small Angle ◽  
Catalytic Domain ◽  
Adaptive Immune Response ◽  
Saxs Data ◽  
Single Stranded Dna ◽  
X Ray ◽  
Adaptive Immune ◽  
X Ray Scattering ◽  
Binding Substrate ◽  
Ray Scattering

In normal cells APOBEC3 (A3A-A3H) enzymes as part of the innate immune system deaminate cytosine to uracil on single-stranded DNA (ssDNA) to scramble DNA in order to give protection against a range of exogenous retroviruses, DNA-based parasites, and endogenous retroelements. However, some viruses and cancer cells use these enzymes, especially A3A and A3B, to escape the adaptive immune response and thereby lead to the evolution of drug resistance. We have synthesized first-in-class inhibitors featuring modified ssDNA. We present models based on small-angle X-ray scattering (SAXS) data that (1) confirm that the mode of binding of inhibitor to an active A3B C-terminal domain construct in the solution state is the same as the mode of binding substrate to inactive mutants of A3A and A3B revealed in X-ray crystal structures and (2) give insight into the disulfide-linked inactive dimer formed under the oxidizing conditions of purification.

Structural refinement by restrained molecular-dynamics algorithm with small-angle X-ray scattering constraints for a biomolecule

2004 ◽  
Vol 37 (1) ◽  
pp. 103-109 ◽  
Author(s):  
Masaki Kojima ◽  
Alexander A. Timchenko ◽  
Junichi Higo ◽  
Kazuki Ito ◽  
Hiroshi Kihara ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Molecular Dynamics ◽  
Small Angle ◽  
Protein Structures ◽  
Saxs Data ◽  
X Ray ◽  
X Ray Scattering ◽  
Saxs Curve ◽  
Restrained Molecular Dynamics ◽  
Ray Scattering

A new algorithm to refine protein structures in solution from small-angle X-ray scattering (SAXS) data was developed based on restrained molecular dynamics (MD). In the method, the sum of squared differences between calculated and observed SAXS intensities was used as a constraint energy function, and the calculation was started from given atomic coordinates, such as those of the crystal. In order to reduce the contribution of the hydration effect to the deviation from the experimental (objective) curve during the dynamics, and purely as an estimate of the efficiency of the algorithm, the calculation was first performed assuming the SAXS curve corresponding to the crystal structure as the objective curve. Next, the calculation was carried out with `real' experimental data, which yielded a structure that satisfied the experimental SAXS curve well. The SAXS data for ribonuclease T1, a single-chain globular protein, were used for the calculation, along with its crystal structure. The results showed that the present algorithm was very effective in the refinement and adjustment of the initial structure so that it could satisfy the objective SAXS data.

Quality control of protein standards for molecular mass determinations by small-angle X-ray scattering

2010 ◽  
Vol 43 (2) ◽  
pp. 237-243 ◽  
Author(s):  
Shuji Akiyama
Keyword(s):  
Quality Control ◽  
Molecular Mass ◽  
Small Angle ◽  
Biological Macromolecules ◽  
Average Deviation ◽  
Saxs Data ◽  
X Ray ◽  
X Ray Scattering ◽  
Standard Quality ◽  
Ray Scattering

Small-angle X-ray scattering (SAXS) is a powerful technique with which to evaluate the size and shape of biological macromolecules in solution. Forward scattering intensity normalized relative to the particle concentration,I(0)/c, is useful as a good measure of molecular mass. A general method for deducing the molecular mass from SAXS data is to determine the ratio ofI(0)/cof a target protein to that of a standard protein with known molecular mass. The accuracy of this interprotein calibration is affected considerably by the monodispersity of the prepared standard, as well as by the precision in estimating its concentration. In the present study, chromatographic fractionation followed by hydrodynamic characterization is proposed as an effective procedure by which to prepare a series of monodispersed protein standards. The estimation of molecular mass within an average deviation of 8% is demonstrated using monodispersed bovine serum albumin as a standard. The present results demonstrate the importance of protein standard quality control in order to take full advantage of interprotein calibration.

scholarly journals Biophysical Characterization of Cancer-Related Carbonic Anhydrase IX

2020 ◽  
Vol 21 (15) ◽  
pp. 5277
Author(s):  
Katarina Koruza ◽  
A. Briana Murray ◽  
Brian P. Mahon ◽  
Jesse B. Hopkins ◽  
Wolfgang Knecht ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Carbonic Anhydrase ◽  
Small Angle ◽  
Catalytic Domain ◽  
Carbonic Anhydrase Ix ◽  
Biophysical Characterization ◽  
X Ray ◽  
Ca Ix ◽  
X Ray Scattering ◽  
Ray Scattering

Upregulation of carbonic anhydrase IX (CA IX) is associated with several aggressive forms of cancer and promotes metastasis. CA IX is normally constitutively expressed at low levels in selective tissues associated with the gastrointestinal tract, but is significantly upregulated upon hypoxia in cancer. CA IX is a multi-domain protein, consisting of a cytoplasmic region, a single-spanning transmembrane helix, an extracellular CA catalytic domain, and a proteoglycan-like (PG) domain. Considering the important role of CA IX in cancer progression and the presence of the unique PG domain, little information about the PG domain is known. Here, we report biophysical characterization studies to further our knowledge of CA IX. We report the 1.5 Å resolution crystal structure of the wild-type catalytic domain of CA IX as well as small angle X-ray scattering and mass spectrometry of the entire extracellular region. We used matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry to characterize the spontaneous degradation of the CA IX PG domain and confirm that it is only the CA IX catalytic domain that forms crystals. Small angle X-ray scattering analysis of the intact protein indicates that the PG domain is not randomly distributed and adopts a compact distribution of shapes in solution. The observed dynamics of the extracellular domain of CA IX could have physiological relevance, including observed cleavage and shedding of the PG domain.

Small-Angle X-Ray Scattering for the Discerning Macromolecular Crystallographer

2014 ◽  
Vol 67 (12) ◽  
pp. 1786 ◽  
Author(s):  
Lachlan W. Casey ◽  
Alan E. Mark ◽  
Bostjan Kobe
Keyword(s):  
Structural Biology ◽  
Small Angle ◽  
Significant Risk ◽  
Macromolecular Crystallography ◽  
Saxs Data ◽  
X Ray ◽  
X Ray Scattering ◽  
Ray Scattering

The role of small-angle X-ray scattering (SAXS) in structural biology is now well established, and its usefulness in combination with macromolecular crystallography is clear. However, the highly averaged SAXS data present a significant risk of over-interpretation to the unwary practitioner, and it can be challenging to frame SAXS results in a manner that maximises the reliability of the conclusions drawn. In this review, a series of recent examples are used to illustrate both the challenges for interpretation and approaches through which these can be overcome.

Influence of polychromaticity on particle structure determination in small-angle X-ray scattering

2015 ◽  
Vol 48 (6) ◽  
pp. 1935-1942 ◽  
Author(s):  
Wenjia Wang ◽  
Eleonora V. Shtykova ◽  
Vladimir V. Volkov ◽  
Guangcai Chang ◽  
Lianhui Zhang ◽  
...  
Keyword(s):  
Small Angle ◽  
Structural Parameters ◽  
Shape Reconstruction ◽  
The Body ◽  
Particle Structure ◽  
Saxs Data ◽  
Time Resolved ◽  
X Ray ◽  
X Ray Scattering ◽  
Ray Scattering

Pink beams are now widely used for small-angle X-ray scattering (SAXS) data collection owing to their high intensity. However, the wavelength spread of a pink beam is a factor of 100 higher than that of a monochromatic beam, thus causing the experimental data to be smeared. To reveal the influence of polychromaticity on shape reconstruction, four geometric bodies (sphere, cube, helix and long cylinder) were used for SAXS data analysis. The results reveal that the influence of polychromaticity on the process of shape reconstruction is significantly more dependent on the geometry of the body than on its size. Scattering objects with smoothed scattering curves can tolerate a higher wavelength spread than those with tortuous curves. It is further demonstrated that the structural parameters calculated from the smeared data sets have little deviation from the ideal ones, which indicates the possibility of using a light source with a greater wavelength spread than a conventional pink beam for special time-resolved SAXS experiments. Finally, it is concluded that SAXS data collected in pink-beam mode can be used directly for structural calculations and model reconstructions without a desmearing procedure.

Silicon photodiode detector for small-angle X-ray scattering

1990 ◽  
Vol 23 (5) ◽  
pp. 430-432 ◽  
Author(s):  
P. R. Jemian ◽  
G. G. Long
Keyword(s):  
Small Angle ◽  
Signal To Noise Ratio ◽  
Scintillation Counter ◽  
Detector Response ◽  
Silicon Photodiode ◽  
Double Crystal ◽  
Saxs Data ◽  
X Ray ◽  
X Ray Scattering ◽  
Ray Scattering

A photodiode X-ray detector was built to measure small-angle X-ray scattering (SAXS) at a synchrotron-radiation source in conjunction with a double-crystal diffractometer SAXS camera at photon energies between 5 and 11 keV. The photodiode detector response in this energy range is linear at photon counting rates up to 1012 photons s−1 and thus it was not necessary to attenuate the monochromatic X-ray beam with calibrated foils. SAXS data taken with a scintillation counter and the photodiode detector are compared, demonstrating marked improvement in counting statistics, rate of data acquisition and signal-to-noise ratio.

scholarly journals Guinier peak analysis for visual and automated inspection of small-angle X-ray scattering data

2016 ◽  
Vol 49 (5) ◽  
pp. 1412-1419 ◽  
Author(s):  
Christopher D. Putnam
Keyword(s):  
Small Angle ◽  
Peak Position ◽  
Scattering Data ◽  
Radius Of Gyration ◽  
Automated Inspection ◽  
Elongation Ratio ◽  
Saxs Data ◽  
X Ray ◽  
X Ray Scattering ◽  
Ray Scattering

The Guinier region in small-angle X-ray scattering (SAXS) defines the radius of gyration,Rg, and the forward scattering intensity,I(0). In Guinier peak analysis (GPA), the plot ofqI(q)versus q2transforms the Guinier region into a characteristic peak for visual and automated inspection of data. Deviations of the peak position from the theoretical position in dimensionless GPA plots can suggest parameter errors, problematic low-resolution data, some kinds of intermolecular interactions or elongated scatters. To facilitate automated analysis by GPA, the elongation ratio (ER), which is the ratio of the areas in the pair-distribution functionP(r) after and before theP(r) maximum, was characterized; symmetric samples have ER values around 1, and samples with ER values greater than 5 tend to be outliers in GPA analysis. Use of GPA+ER can be a helpful addition to SAXS data analysis pipelines.

scholarly journals Scaling Behavior of Single Stranded DNA Measured by Small Angle X-ray Scattering

2009 ◽  
Vol 96 (3) ◽  
pp. 347a
Author(s):  
Adelene Y.L. Sim ◽  
Jan Lipfert ◽  
Daniel Herschlag ◽  
Sebastian Doniach
Keyword(s):  
Small Angle ◽  
Scaling Behavior ◽  
Single Stranded Dna ◽  
X Ray ◽  
X Ray Scattering ◽  
Ray Scattering

scholarly journals Obtaining tertiary protein structures by the ab-initio interpretation of small angle X-ray scattering data

2019 ◽  
Author(s):  
Christopher Prior ◽  
Owen R Davies ◽  
Daniel Bruce ◽  
Ehmke Pohl
Keyword(s):  
Ab Initio ◽  
Small Angle ◽  
Protein Structures ◽  
Scattering Data ◽  
Ab Initio Method ◽  
Primary Sequence ◽  
Saxs Data ◽  
X Ray ◽  
X Ray Scattering ◽  
Ray Scattering

ABSTRACTSmall angle X-ray scattering (SAXS) has become an important tool to investigate the structure of proteins in solution. In this paper we present a novel ab-initio method to represent polypeptide chains as discrete curves that can be used to derive a meaningful three-dimensional model from only the primary sequence and experimental SAXS data. High resolution crystal structures were used to generate probability density functions for each of the common secondary structural elements found in proteins. These are used to place realistic restraints on the model curve’s geometry. To evaluate the quality of potential models and demonstrate the efficacy of this novel technique we developed a new statistic to compare the entangled geometry of two open curves, based on mathematical techniques from knot theory. The chain model is coupled with a novel explicit hydration shell model in order derive physically meaningful 3D models by optimizing configurations against experimental SAXS data using a monte-caro based algorithm. We show that the combination of our ab-initio method with spatial restraints based on contact predictions successfully derives a biologically plausible model of the coiled–coil component of the human synaptonemal complex central element protein.SIGNIFICANCESmall-angle X-ray scattering allows for structure determination of biological macromolecules and their complexes in aqueous solution. Using a discrete curve representation of the polypeptide chain and combining it with empirically determined constraints and a realistic solvent model we are now able to derive realistic ab-initio 3-dimensional models from BioSAXS data. The method only require a primary sequence and the scattering data form the user.

scholarly journals Structure of Nonionic Surfactant Diglycerol Monomyristate Micelles in Cyclohexane: a SAXS Study

1970 ◽  
Vol 23 ◽  
pp. 74-81
Author(s):  
Lok Kumar Shrestha
Keyword(s):  
Nonionic Surfactant ◽  
Small Angle ◽  
Water Addition ◽  
Saxs Data ◽  
Cross Sectional ◽  
X Ray ◽  
X Ray Scattering ◽  
Added Water ◽  
Trace Water ◽  
Ray Scattering

Structure of nonionic surfactant diglycerol monomyristate (C14G2) micelles in cyclohexane has been investigated by small-angle X-ray scattering (SAXS) technique. Structural modulation of reverse micelle (RM) has been systematically studied by changing composition, temperature change and added-water. The SAXS data were evaluated by the generalized indirect Fourier transformation (GIFT) method, which gives pair-distance distribution function (PDDF). Unlike conventional poly(oxyethylene) type nonionic surfactants, C14G2 forms RM in cyclohexane without water addition at normal room temperature. A clear indication of one dimensional (1-D) micellar growth was found with increasing C14G2 concentrations. On the other hand, temperature induced cylinder-to-sphere type transition in the RM structure. The maximum dimension and the cross-sectional diameter of the RM increased upon addition of trace water indicating the formation of water pool in the reverse micellar core.Keywords: Diglycerol monomyristate, small-angle X-ray scattering, reverse micelles.DOI: 10.3126/jncs.v23i0.2099J. Nepal Chem. Soc., Vol. 23, 2008/2009Page: 74-81

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