title: Dissecting the effect of RNA aptamer binding on the dynamics of plasminogen activator inhibitor 1 using hydrogen/deuterium exchange mass spectrometry.
abstract: RNA aptamers, selected from large synthetic libraries, are attracting increasing interest as protein ligands, with potential uses as prototype pharmaceuticals, conformational probes, and reagents for specific quantification of protein levels in biological samples. Very little is known, however, about their effects on protein conformation and dynamics. We have employed hydrogen/deuterium exchange (HDX) mass spectrometry to study the effect of RNA aptamers on the structural flexibility of the serpin plasminogen activator inhibitor-1 (PAI-1). The aptamers have characteristic effects on the biochemical properties of PAI-1. In particular, they are potent inhibitors of the structural transition of PAI-1 from the active state to the inactive, so-called latent state. This transition is one of the largest conformational changes of a folded protein domain without covalent modification. Binding of the aptamers to PAI-1 is associated with substantial and widespread protection against deuterium uptake in PAI-1. The aptamers induce protection against exchange with the solvent both in the protein-aptamer interface as well as in other specific areas. Interestingly, the aptamers induce substantial protection against exchange in α-helices B, C and I. This observation substantiates the relevance of structural instability in this region for transition to the latent state and argues for involvement of flexibility in regions not commonly associated with regulation of latency transition in serpins.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Southern Denmark , Campusvej 55, DK-5230 Odense M, Denmark.
authors.forename: Morten B
authors.initials: MB
authors.lastname: Trelle
authors.forename: Daniel M
authors.initials: DM
authors.lastname: Dupont
authors.forename: Jeppe B
authors.initials: JB
authors.lastname: Madsen
authors.forename: Peter A
authors.initials: PA
authors.lastname: Andreasen
authors.forename: Thomas J D
authors.initials: TJ
authors.lastname: Jørgensen
mesh_terms: D052157:Aptamers, Nucleotide / Q000378:metabolism*; D003903:Deuterium / Q000737:chemistry; D006801:Humans; D006859:Hydrogen / Q000737:chemistry; D013058:Mass Spectrometry; D008958:Models, Molecular; D017395:Plasminogen Activator Inhibitor 1 / Q000737:chemistry* / Q000378:metabolism*; D011487:Protein Conformation; D055550:Protein Stability
chemical_list: D052157:Aptamers, Nucleotide; D017395:Plasminogen Activator Inhibitor 1; D006859:Hydrogen; D003903:Deuterium