A modular system for sequence-specific DNA recognition in the major groove

• Applicant: Professor Dr. Thomas Schrader
• Subject Area: Organic Molecular Chemistry - Synthesis and Characterisation ; Biological and Biomimetic Chemistry
• Term: from 2015 to 2019
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 271356290

Nucleic acid recognition is a fundamental biological process. For the purpose of adressing a specific gene, a natural or artificial binding partner must be directed towards the minor or major groove, because here the base sequence becomes freely accessible - and readable. In the past decades, sophisticated synthetic minor and major groove binders have been created. However, while for the minor groove a complete set of oligoamides has been developed which allows specific targeting of any small base sequence, such a universal solution has been elusive for DNAs major groove.Triplex forming oligonucleotides (TFOs) are largely limited to homopurines; and although a number of alternative nucleic acid skeletons have been designed (PNA, LNA), most of these do not preserve the intact duplex, but invade and open the existing double strand. Despite several attempts, there is until today, a frustrating lack of entirely artificial non-invasive sequence-selective major groove binders for ds-DNA which operate efficiently without sequence constraints.In this proposal we want to delineate the development of such a general modular set of recognition units for DNA double strands, which operate by triplex formation inside the major groove. Each module contains a powerful base pair binder attached to a PNA element. All available donors and acceptors in the major groove are thus hydrogen-bonded by the base pair binders; further stabilization comes from attractive interactions with the phosphodiester backbone as well as from extensive pi-stacking among the heterocyclic ring systems. Covalent module connection with a consecutive arrangement of appropriate base pair binders leads to a new PNA derivative which is perfectly complementary to a selected short ds DNA fragment. Thus, only 4 different modules (AT-, TA-, CG-, GC-binder) suffice for the iterative coupling on a standard peptide synthesizer. The construction of synthetic sequence-selective DNA ligands which operate by general base-pair recognition inside the major groove represents a formidable challenge for supramolecular chemists; in this proposal, the emphasis lies on the construction, coupling and DNA binding properties of the new modules involving in-vitro binding studies with isolated oligonucleotides. A second project stage (another 3 years) is envisioned, which will apply the optimized modules in vivo (cell culture and, if possible, animals). Deliberate interference with specific DNA fragments can be used for site-specific modulation of gene expression, modulation of protein binding, targeting of DNA damage, mutagenesis and enhancement of homologous recombination, thus providing a tool for gene-specific manipulation of DNA. Such new TFOs may be used, on the other hand to deliver cleaving or cross-linking agents, transcription factors or nucleases to a chosen site on the DNA, both in vitro and in vivo.

• DFG Programme: Research Grants