Spectral and Hydrodynamic Analysis of West Nile Virus RNA-Protein Interactions by Multiwavelength Sedimentation Velocity in the Analytical Ultracentrifuge.
Anal Chem
; 89(1): 862-870, 2017 01 03.
Article
em En
| MEDLINE
| ID: mdl-27977168
ABSTRACT
Interactions between nucleic acids and proteins are critical for many cellular processes, and their study is of utmost importance to many areas of biochemistry, cellular biology, and virology. Here, we introduce a new analytical method based on sedimentation velocity (SV) analytical ultracentrifugation, in combination with a novel multiwavelength detector to characterize such interactions. We identified the stoichiometry and molar mass of a complex formed during the interaction of a West Nile virus RNA stem loop structure with the human T cell-restricted intracellular antigen-1 related protein. SV has long been proven as a powerful technique for studying dynamic assembly processes under physiological conditions in solution. Here, we demonstrate, for the first time, how the new multiwavelength technology can be exploited to study protein-RNA interactions, and show how the spectral information derived from the new detector complements the traditional hydrodynamic information from analytical ultracentrifugation. Our method allows the protein and nucleic acid signals to be separated by spectral decomposition such that sedimentation information from each individual species, including any complexes, can be clearly identified based on their spectral signatures. The method presented here extends to any interacting system where the interaction partners are spectrally separable.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Ultracentrifugação
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Vírus do Nilo Ocidental
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RNA Viral
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Hidrodinâmica
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Antígeno-1 Intracelular de Células T
Limite:
Humans
Idioma:
En
Revista:
Anal Chem
Ano de publicação:
2017
Tipo de documento:
Article
País de afiliação:
Estados Unidos