Small-angle scattering contrast calculator for protein and nucleic acid complexes in solution

Sarachan, Kathryn L.; Curtis, Joseph E.; Krueger, Susan

doi:10.1107/s0021889813025727

Cited by 34 publications

(24 citation statements)

References 14 publications

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“…8) to further confirm the R g and I (0) values. Theoretical Δ ρ 1 and Δ ρ 2 values were calculated for the Skp and uOMP components and theoretical

\sqrt{\frac{true}{I false(0 false)}}

values were calculated for the complexes as a function of f D2O from the protein sequences using the Contrast Calculator (Sarachan et al, 2013) module of the SASSIE (Curtis et al, 2012) software assuming different percentages of deuteration for the OmpW and OmpA. The

\sqrt{\frac{true}{I false(0 false)}}

values were compared to those obtained from the Guinier analysis, allowing for experimental verification of the complex match point and the amount of deuteration of the uOMP component in the measured samples.…”

Section: Materials and Methods*mentioning

confidence: 99%

“…5), the contrast match point of the complex can be determined from the x-intercept of a linear fit to

\sqrt{\frac{true}{I false(0 false)}}

versus f D2O . The match point of the complex can also be calculated from the sequences of the components (Sarachan, Curtis, & Krueger, 2013; Whitten, Cai, & Trewhella, 2008). Using both approaches to determine the match point provides another quality assurance test on the data in that the calculated and experimentally determined match points should agree with each other.…”

Section: Sans From Biological Molecules In Solutionmentioning

confidence: 99%

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Deuterium Labeling Together with Contrast Variation Small-Angle Neutron Scattering Suggests How Skp Captures and Releases Unfolded Outer Membrane Proteins

Zaccai

Sandlin

Hoopes

et al. 2016

Methods in Enzymology

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In gram-negative bacteria, the chaperone protein Skp forms specific and stable complexes with membrane proteins while they are transported across the periplasm to the outer membrane. The jellyfish-like architecture of Skp is similar to the eukaryotic and archeal prefoldins and the mitochondrial Tim chaperones, that is α-helical ‘tentacles’ extend from a β-strand ‘body’ to create an internal cavity. Contrast variation small-angle neutron scattering (SANS) experiments on Skp alone in solution and bound in two different complexes to unfolded outer membrane proteins (uOMPs), OmpA and OmpW, demonstrate that the helical tentacles of Skp bind their substrate in a clamp-like mechanism in a conformation similar to that previously observed in the apo crystal structure of Skp. Deuteration of the uOMP component combined with contrast variation analysis allowed the shapes of Skp and uOMP as well as the location of uOMP with respect to Skp to be determined in both complexes. This represents unique information that could not be obtained without deuterium labeling of the uOMPs. The data yield the first direct structural evidence that the α-helical Skp tentacles move closer together on binding its substrate and that the structure of Skp is different when binding different uOMPs. This work presents, by example, a tutorial on performing SANS experiments using both deuterium labeling and contrast variation, including SANS theory, sample preparation, data collection, sample quality validation, data analysis and structure modeling.

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“…8) to further confirm the R g and I (0) values. Theoretical Δ ρ 1 and Δ ρ 2 values were calculated for the Skp and uOMP components and theoretical

\sqrt{\frac{true}{I false(0 false)}}

\sqrt{\frac{true}{I false(0 false)}}

Section: Materials and Methods*mentioning

confidence: 99%

“…5), the contrast match point of the complex can be determined from the x-intercept of a linear fit to

\sqrt{\frac{true}{I false(0 false)}}

Section: Sans From Biological Molecules In Solutionmentioning

confidence: 99%

Deuterium Labeling Together with Contrast Variation Small-Angle Neutron Scattering Suggests How Skp Captures and Releases Unfolded Outer Membrane Proteins

Zaccai

Sandlin

Hoopes

et al. 2016

Methods in Enzymology

Self Cite

View full text Add to dashboard Cite

show abstract

“…where Á M is the change in scattering length per mass, Á M = ð M;protein À solvent Þ " r 0 , where " is the protein partial specific volume (set to 0.7425 cm 3 g À1 ) and r 0 is the scattering length of an electron (Mylonas & Svergun, 2007). M,protein of rHSA was estimated to be 428 electrons nm À3 , as described by Sarachan et al (2013). N A is Avogrado's constant and MM is the molecular mass.…”

Section: Analysis and Scaling Of Datamentioning

confidence: 99%

Concentrated protein solutions investigated using acoustic levitation and small-angle X-ray scattering

Sønderby

Söderberg

Frankær

et al. 2020

J Synchrotron Radiat

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An acoustically levitated droplet has been used to collect synchrotron SAXS data on human serum albumin protein solutions up to a protein concentration of 400 mg ml−1. A careful selection of experiments allows for fast data collection of a large amount of data, spanning a protein concentration/solvent concentration space with limited sample consumption (down to 3 µL per experiment) and few measurements. The data analysis shows data of high quality that are reproducible and comparable with data from standard flow‐through capillary‐based experiments. Furthermore, using this methodology, it is possible to achieve concentrations that would not be accessible by conventional cells. The protein concentration and ionic strength parameter space diagram may be covered easily and the amount of protein sample is significantly reduced (by a factor of 100 in this work). Used in routine measurements, the benefits in terms of protein cost and time spent are very significant.

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“…Provided with the chemical composition and volumes of small molecules or sequences in the case of biomacromolecules (proteins and RNA/DNA), it calculates contrast, R g and interference terms as a function of buffer conditions (H 2 O/D 2 O ratio, the presence of small cosolutes such as salt etc.). The SASSIE contrast calculator SCC (Sarachan et al, 2013) is an integrated module of the SASSIE suite (Curtis et al, 2012). It calculates contrast, contrast-match points (CMPs) and forward scattering intensities [I(0)] in absolute units for proteins, nucleic acids (RNA/DNA) and small molecules (lipids/detergents) and their complexes based on their primary sequences or chemical composition, deuteration level and concentration.…”

Section: Software Developmentsmentioning

confidence: 99%

Biological small-angle neutron scattering: recent results and development

Mahieu

Gabel

2018

Acta Cryst Sect D Struct Biol

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Small-angle neutron scattering (SANS) has increasingly been used by the structural biology community in recent years to obtain low-resolution information on solubilized biomacromolecular complexes in solution. In combination with deuterium labelling and solvent-contrast variation (HO/DO exchange), SANS provides unique information on individual components in large heterogeneous complexes that is perfectly complementary to the structural restraints provided by crystallography, nuclear magnetic resonance and electron microscopy. Typical systems studied include multi-protein or protein-DNA/RNA complexes and solubilized membrane proteins. The internal features of these systems are less accessible to the more broadly used small-angle X-ray scattering (SAXS) technique owing to a limited range of intra-complex and solvent electron-density variation. Here, the progress and developments of biological applications of SANS in the past decade are reviewed. The review covers scientific results from selected biological systems, including protein-protein complexes, protein-RNA/DNA complexes and membrane proteins. Moreover, an overview of recent developments in instruments, sample environment, deuterium labelling and software is presented. Finally, the perspectives for biological SANS in the context of integrated structural biology approaches are discussed.

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Small-angle scattering contrast calculator for protein and nucleic acid complexes in solution

Cited by 34 publications

References 14 publications

Deuterium Labeling Together with Contrast Variation Small-Angle Neutron Scattering Suggests How Skp Captures and Releases Unfolded Outer Membrane Proteins

Deuterium Labeling Together with Contrast Variation Small-Angle Neutron Scattering Suggests How Skp Captures and Releases Unfolded Outer Membrane Proteins

Concentrated protein solutions investigated using acoustic levitation and small-angle X-ray scattering

Biological small-angle neutron scattering: recent results and development

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