Size-exclusion chromatography small-angle X-ray scattering of water soluble proteins on a laboratory instrument

Bucciarelli, Saskia; Midtgaard, Søren Roi; Pedersen, Martin Nors; Skou, Søren; Arleth, Lise; Vestergaard, Bente

doi:10.1107/s1600576718014462

Cited by 40 publications

(27 citation statements)

References 64 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…In many cases, SEC-SAXS can effectively separate scattering from aggregates and single proteins. Recently, this option also became available for SANS (SEC-SANS; Jordan et al, 2016) and for home-source SAXS instruments (Bucciarelli et al, 2018). However, when the amount of protein is limited, and several conditions such as temperature, pH and concentration dependence need to be tested, then SEC-SAXS and SEC-SANS may not be practically feasible.…”

Section: Detecting and Reducing Undesired Aggregation In Saxs And Sanmentioning

confidence: 99%

Assessment of structure factors for analysis of small-angle scattering data from desired or undesired aggregates

Larsen¹,

Pedersen²,

Arleth³

2020

J Appl Cryst

Self Cite

View full text Add to dashboard Cite

Aggregation processes are central features of many systems ranging from colloids and polymers to inorganic nanoparticles and biological systems. Some aggregated structures are controlled and desirable, e.g. in the design of size-controlled clustered nanoparticles or some protein-based drugs. In other cases, the aggregates are undesirable, e.g. protein aggregation involved in neurodegenerative diseases or in vitro studies of single protein structures. In either case, experimental and analytical tools are needed to cast light on the aggregation processes. Aggregation processes can be studied with small-angle scattering, but analytical descriptions of the aggregates are needed for detailed structural analysis. This paper presents a list of useful small-angle scattering structure factors, including a novel structure factor for a spherical cluster with local correlations between the constituent particles. Several of the structure factors were renormalized to get correct limit values in both the high-q and low-q limit, where q is the modulus of the scattering vector. The structure factors were critically evaluated against simulated data. Structure factors describing fractal aggregates provided approximate descriptions of the simulated data for all tested structures, from linear to globular aggregates. The addition of a correlation hole for the constituent particles in the fractal structure factors significantly improved the fits in all cases. Linear aggregates were best described by a linear structure factor and globular aggregates by the newly derived spherical cluster structure factor. As a central point, it is shown that the structure factors could be used to take aggregation contributions into account for samples of monomeric protein containing a minor fraction of aggregated protein. After applying structure factors in the analysis, the correct structure and oligomeric state of the protein were determined. Thus, by careful use of the presented structure factors, important structural information can be retrieved from small-angle scattering data, both when aggregates are desired and when they are undesired.

show abstract

Section: Detecting and Reducing Undesired Aggregation In Saxs And Sanmentioning

confidence: 99%

Assessment of structure factors for analysis of small-angle scattering data from desired or undesired aggregates

Larsen¹,

Pedersen²,

Arleth³

2020

J Appl Cryst

Self Cite

View full text Add to dashboard Cite

show abstract

“…Também pode ser empregada no fracionamento de substâncias húmicas em diferentes sistemas de cultivo e em diferentes matrizes, para determinar parâmetros moleculares como o raio hidrodinâmico e a comum massa molecular, a distribuição do peso molecular de metais nos extratos de biomassa de fungos ou extrair e separar com sucesso diferentes frações de peso molecular de diferentes matrizes. [68][69][70][71][72][73][74][75][76] A cromatografia de exclusão por tamanho também pode ser acoplada a um espectrômetro de massa com plasma indutivamente acoplado, tornandose uma técnica eficiente de detecção e especiação de metaloproteínas ou metalobiomoléculas. 77 Como pode ser visto no trabalho de Lopes Júnior et al 77 em que buscaram avaliar a influência do cádmio no metabolismo do girassol, os autores utilizaram a SEC-ICP-MS para detectar a presença de metalobiomoléculas nas sementes de girassol e observaram que esse metal se ligava normalmente a compostos de massas moleculares de 3 a 70 kDa.…”

Section: Cromatografia Por Exclusão De Tamanhosunclassified

Metallomics: Methodologies and Applications in Environmental Studies

Silva¹,

Silva²

2020

Rev. Virtual Quim.

View full text Add to dashboard Cite

Metallomics is an interdisciplinary area that has grown in recent years thanks to the development and application of techniques such as electrophoresis, chromatography, and spectrometry. Advances in this area, have allowed studies of functional and structural characterization of proteins linked to metals in different organisms and tissues, and thus, led to the understanding of the complex processes involving metalloproteins, and with this, their application for the realization of environmental monitoring studies. In this article, a review of the main metallomic techniques and their applications in environmental studies was carried out from 2008 to 2018, based on the search for scientific articles in the "Scopus" database. A metalômica é uma área interdisciplinar que tem crescido nos últimos anos, em virtude ao desenvolvimento e aplicação de técnicas como eletroforese, cromatografia e espectrometria. Avanços nessa área, têm permitido estudos de caracterização funcional e estrutural de proteínas ligadas a metais em diferentes organismos e tecidos, e desta forma, levado à compreensão, dos complexos processos que envolvem as metaloproteínas, e com isto, garantido sua aplicação para a realização de estudos de monitoramento ambiental. No presente artigo, foi realizada uma revisão das principais técnicas metalômicas e de suas aplicações em estudos ambientais no período de 2008 a 2018, a partir da busca de artigos científicos na base de dados "Scopus".

show abstract

“…Under this situation, Bucciarelli et al . 8 launched a laboratory-based SEC-SAXS system and succeeded to obtain the SAXS data, which enables the further analysis of a non-aggregated target component for the first time. Here, at least three technical improvements must be considered for a next generation laboratory-based SEC-SAXS.…”

Section: Introductionmentioning

confidence: 99%

Newly developed Laboratory-based Size exclusion chromatography Small-angle x-ray scattering System (La-SSS)

Inoue

Nakagawa²,

Morishima

et al. 2019

Sci Rep

View full text Add to dashboard Cite

To understand a biological system, it is important to observe structures of biomolecules in the solution where the system is functionalized. S mall- A ngle X -ray S cattering coupled with S ize E xclusion C hromatography (SEC-SAXS) is one of techniques to selectively observe the target molecules in the multi-component system. However, this technique is believed to be available only with a synchrotron-based SAXS instrument due to requirement of high beam intensity and, therefore, the limitation of the beam time was obstacle to satisfy demands from many bio-researchers. We newly developed La boratory-based S ize exclusion chromatography S AXS S ystem (La-SSS) by utilizing a latest laboratory-based SAXS instrument and finely optimization of the balance between flow rate, cell volume, irradiation time and so on. La-SSS succeeded not only decoupling of target protein(s) from non-specific aggregates but also measurement of each concerned component in a multi-component system. In addition, an option: “stopping mode”, which is designed for improving statistics of SAXS profile, realized a high S/N data acquisition for the most interesting protein in a multi-component system. Furthermore, by utilizing a column having small bed volume, the small-scale SEC-SAXS study makes available. Through optimization of instrumental parameters and environments, La-SSS is highly applicable for experimental requirements from various biological samples. It is strongly expected that a La-SSS concept must be a normal option for laboratory-based SAXS in the near future.

show abstract

Size-exclusion chromatography small-angle X-ray scattering of water soluble proteins on a laboratory instrument

Abstract: Size-exclusion chromatography coupled with a laboratory-based small-angle X-ray scattering setup is presented and its performance evaluated for studies of various proteins covering a broad range of molecular weights.

Cited by 40 publications

References 64 publications

Assessment of structure factors for analysis of small-angle scattering data from desired or undesired aggregates

Assessment of structure factors for analysis of small-angle scattering data from desired or undesired aggregates

Metallomics: Methodologies and Applications in Environmental Studies

Newly developed Laboratory-based Size exclusion chromatography Small-angle x-ray scattering System (La-SSS)

Contact Info

Product

Resources

About