Toehold-Mediated Strand Displacement in a Triplex Forming Nucleic Acid Clamp for Reversible Regulation of Polymerase Activity and Protein Expression

TY - JOUR

T1 - Toehold-Mediated Strand Displacement in a Triplex Forming Nucleic Acid Clamp for Reversible Regulation of Polymerase Activity and Protein Expression

AU - Nguyen, Thuy J.D.

AU - Manuguerra, Ilenia

AU - Kumar, Vipin

AU - Gothelf, Kurt V.

PY - 2019

Y1 - 2019

N2 - Triplex forming oligonucleotides are used as a tool for gene regulation and in DNA nanotechnology. By incorporating artificial nucleic acids, target affinity and biological stability superior to that of natural DNA may be obtained. This work demonstrates how a chimeric clamp consisting of acyclic (L)-threoninol nucleic acid (aTNA) and DNA can bind DNA and RNA by the formation of a highly stable triplex structure. The (L)-aTNA clamp is released from the target again by the addition of a releasing strand in a strand displacement type of reaction. It is shown that the clamp efficiently inhibits Bsu and T7 RNA polymerase activity and that polymerase activity is reactivated by displacing the clamp. The clamp was successfully applied to the regulation of luciferase expression by reversible binding to the mRNA. When targeting a sequence in the double stranded plasmid, 40 % downregulation of protein expression is achieved.

AB - Triplex forming oligonucleotides are used as a tool for gene regulation and in DNA nanotechnology. By incorporating artificial nucleic acids, target affinity and biological stability superior to that of natural DNA may be obtained. This work demonstrates how a chimeric clamp consisting of acyclic (L)-threoninol nucleic acid (aTNA) and DNA can bind DNA and RNA by the formation of a highly stable triplex structure. The (L)-aTNA clamp is released from the target again by the addition of a releasing strand in a strand displacement type of reaction. It is shown that the clamp efficiently inhibits Bsu and T7 RNA polymerase activity and that polymerase activity is reactivated by displacing the clamp. The clamp was successfully applied to the regulation of luciferase expression by reversible binding to the mRNA. When targeting a sequence in the double stranded plasmid, 40 % downregulation of protein expression is achieved.

KW - gene knockdown

KW - nucleic acids

KW - primer extension

KW - protein expression

KW - threoninol

KW - triplex formation

UR - https://www.scopus.com/pages/publications/85071339386

U2 - 10.1002/chem.201903496

DO - 10.1002/chem.201903496

M3 - Journal article

C2 - 31373735

AN - SCOPUS:85071339386

SN - 0947-6539

VL - 25

SP - 12303

EP - 12307

JO - Chemistry - A European Journal

JF - Chemistry - A European Journal

IS - 53

ER -