title: Building a Molecular Trap for a Serine Protease from Aptamer and Peptide Modules.
abstract: In drug development, molecular intervention strategies are usually based on interference with a single protein function, such as enzyme activity or receptor binding. However, in many cases, protein drug targets are multifunctional, with several molecular functions contributing to their pathophysiological actions. Aptamers and peptides are interesting synthetic building blocks for the design of multivalent molecules capable of modulating multiple functions of a target protein. Here, we report a molecular trap with the ability to interfere with the activation, catalytic activity, receptor binding, etc. of the serine protease urokinase-type plasminogen activator (uPA) by a rational combination of two RNA aptamers and a peptide with different inhibitory properties. The assembly of these artificial inhibitors into one molecule enhanced the inhibitory activity between 10- and 10,000-fold toward several functions of uPA. The study highlights the potential of multivalent designs and illustrates how they can easily be constructed from aptamers and peptides using nucleic acid engineering, chemical synthesis, and bioconjugation chemistry. By aptamer to aptamer and aptamer to peptide conjugation, we created, to the best of our knowledge, the first trivalent molecule which combines three artificial inhibitors binding to three different sites in a protein target. We hypothesize that by simultaneously preventing all of the functional interactions and activities of the target protein, this approach may represent an alternative to siRNA technology for a functional knockout.
authors.affiliation: Department of Molecular Biology and Genetics, Aarhus University , Gustav Wieds Vej 10C, 8000 Aarhus, Denmark.
authors.forename: Daniel M
authors.initials: DM
authors.lastname: Dupont
authors.affiliation: Department of Molecular Biology and Genetics, Aarhus University , Gustav Wieds Vej 10C, 8000 Aarhus, Denmark.
authors.forename: Nils
authors.initials: N
authors.lastname: Bjerregaard
authors.affiliation: Department of Molecular Biology and Genetics, Aarhus University , Gustav Wieds Vej 10C, 8000 Aarhus, Denmark.
authors.forename: Ben
authors.initials: B
authors.lastname: Verpaalen
authors.affiliation: Department of Molecular Biology and Genetics, Aarhus University , Gustav Wieds Vej 10C, 8000 Aarhus, Denmark.
authors.forename: Peter A
authors.initials: PA
authors.lastname: Andreasen
authors.affiliation: Department of Molecular Biology and Genetics, Aarhus University , Gustav Wieds Vej 10C, 8000 Aarhus, Denmark.
authors.forename: Jan K
authors.initials: JK
authors.lastname: Jensen
mesh_terms: D000595:Amino Acid Sequence; D052157:Aptamers, Nucleotide / Q000737:chemistry*; D010455:Peptides / Q000737:chemistry*; D057057:Serine Proteases / Q000737:chemistry*
chemical_list: D052157:Aptamers, Nucleotide; D010455:Peptides; D057057:Serine Proteases