title: Protein-RNA specificity by high-throughput principal component analysis of NMR spectra.
abstract: Defining the RNA target selectivity of the proteins regulating mRNA metabolism is a key issue in RNA biology. Here we present a novel use of principal component analysis (PCA) to extract the RNA sequence preference of RNA binding proteins. We show that PCA can be used to compare the changes in the nuclear magnetic resonance (NMR) spectrum of a protein upon binding a set of quasi-degenerate RNAs and define the nucleobase specificity. We couple this application of PCA to an automated NMR spectra recording and processing protocol and obtain an unbiased and high-throughput NMR method for the analysis of nucleobase preference in protein-RNA interactions. We test the method on the RNA binding domains of three important regulators of RNA metabolism.
authors.affiliation: Molecular Structure Division, MRC National Institute for Medical Research, London NW7 1AA, UK Research Department of Structural and Molecular Biology, University College London, London WC1E 6BT, UK.
authors.forename: Katherine M
authors.initials: KM
authors.lastname: Collins
authors.affiliation: MRC Biomedical NMR Centre, MRC National Institute for Medical Research, London NW7 1AA, UK.
authors.forename: Alain
authors.initials: A
authors.lastname: Oregioni
authors.affiliation: Molecular Structure Division, MRC National Institute for Medical Research, London NW7 1AA, UK.
authors.forename: Laura E
authors.initials: LE
authors.lastname: Robertson
authors.affiliation: MRC Biomedical NMR Centre, MRC National Institute for Medical Research, London NW7 1AA, UK a.ramos@ucl.ac.uk.
authors.forename: Geoff
authors.initials: G
authors.lastname: Kelly
authors.affiliation: Molecular Structure Division, MRC National Institute for Medical Research, London NW7 1AA, UK Research Department of Structural and Molecular Biology, University College London, London WC1E 6BT, UK a.ramos@ucl.ac.uk.
authors.forename: Andres
authors.initials: A
authors.lastname: Ramos
mesh_terms: D001483:Base Sequence; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D057166:High-Throughput Screening Assays / Q000379:methods* / Q000706:statistics & numerical data; D006801:Humans; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D025341:Principal Component Analysis; D054730:Protein Interaction Domains and Motifs; D012313:RNA / Q000235:genetics* / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism; D039221:mRNA Cleavage and Polyadenylation Factors / Q000737:chemistry / Q000378:metabolism
chemical_list: D004268:DNA-Binding Proteins; C118777:KHDRBS3 protein, human; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; C511130:TUT4 protein, human; D039221:mRNA Cleavage and Polyadenylation Factors; C512635:RNA15 protein, S cerevisiae; D012313:RNA