title: Probing RegA/RNA interactions using electrospray ionization-fourier transform ion cyclotron resonance-mass spectrometry.
abstract: The interactions of bacteriophage T4 regA protein, a unique translational regulator, with RNAs of various size and sequence were studied using electrospray ionization-Fourier transform ion cyclotron resonance-mass spectrometry. Using very gentle interface conditions, regA/RNA complexes with a 1:1 binding stoichiometry were observed for all four target RNAs studied, consistent with solution binding studies. Competitive binding of target RNAs and their degradation products with regA demonstrated that the loss of a single nucleotide resulted in a dramatic change in binding affinity in some cases. Competitive binding of regA with four target RNAs revealed similar relative binding affinity order to that suggested by previous in vitro repression experiments. The use of sustained off-resonance irradiation for collisionally induced dissociation of a regA/RNA complex suggested the potential for directly obtaining information regarding the regA binding domain.
authors.affiliation: Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory, Richland, Washington, 99352, USA.
authors.forename: C
authors.initials: C
authors.lastname: Liu
authors.forename: L P
authors.initials: LP
authors.lastname: Tolić
authors.forename: S A
authors.initials: SA
authors.lastname: Hofstadler
authors.forename: A C
authors.initials: AC
authors.lastname: Harms
authors.forename: R D
authors.initials: RD
authors.lastname: Smith
authors.forename: C
authors.initials: C
authors.lastname: Kang
authors.forename: N
authors.initials: N
authors.lastname: Sinha
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry*; D001665:Binding Sites / Q000502:physiology; D001667:Binding, Competitive; D017357:Cyclotrons; D013058:Mass Spectrometry / Q000379:methods*; D010455:Peptides / Q000737:chemistry; D012313:RNA / Q000737:chemistry*; D016601:RNA-Binding Proteins / Q000378:metabolism; D014764:Viral Proteins / Q000737:chemistry
chemical_list: D001426:Bacterial Proteins; D010455:Peptides; D016601:RNA-Binding Proteins; C072722:RegA protein, Bacteria; D014764:Viral Proteins; D012313:RNA