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Unraveling nanosecond motions in nucleic acids with high-resolution relaxometry: how dynamic is nicked DNA?

Projektbeschreibung

Bewegungsanalysen von DNA-Molekülen auf atomarer Ebene

Neuere Forschungen gehen bei DNA-Molekülen von einer konformativen Flexibilität aus, die von großer Bedeutung ist, da die DNA mit spezifischen Proteinen interagiert. Diese Flexibilität ist wesentlich von der DNA-Sequenz abhängig und ein entscheidender Faktor bei der Genexpression. Wie flexibel DNA auf Läsionen wie Doppelstrangbrüche reagiert, wurde bislang jedoch nicht erforscht. So untersucht das EU-finanzierte Projekt HRRinDNAwithSSB die Dynamik der DNA-Flexibilität und Bewegungen auf Nanoebene, die diese Eigenschaft beeinflussen. Die Ergebnisse werden Aufschluss über die Bewegungen und Flexibilität geschädigter DNA und deren Reparatur liefern.

Ziel

What role do conformational dynamics play in DNA function and repair? Structures of DNA show local dynamics, conformational flexibility of bases, and large conformational changes in the double helix, indicating easily accessible motions. Yet studying fast motions in nucleic acids is challenging. To address this we will introduce High Resolution Relaxometry (HRR) and apply it to study single strand breaks (SSBs) in DNA. Nucleic acids are often studied at atomic resolution with X-ray crystallography and high-field Nuclear Magnetic Resonance (NMR). Yet neither is suitable to study ns-motions. X-ray crystallography does not report on dynamics while using high-field NMR leads to high resonance frequencies so little ns time-scale information is present. This presents a challenge: how to characterise fast motions in nucleic acids? We will develop a new methodological approach, HRR, to probe ns-motions in DNA. HRR was developed by the host team to study ns-motions in proteins. We will adapt these methods to investigate motions in DNA. We will compare dynamics occurring in intact DNA, DNA with a SSB and SSB DNA with a missing base. Understanding the motions in each DNA construct will establish the effects that each type of DNA damage have on the motional properties of DNA. This will elucidate how each type of damage affects the base pair stacking and the motions occurring at the breakpoint. Understanding the flexibility induced by DNA damage will have a significant role in understanding DNA repair and how damaged DNA is recognised. The DNA repair protein, PARP-1, is a cancer-drug target and recognizes SSBs. Our final objective is to uncover the role of DNA motions in SSB recognition by PARP-1. In summary we will develop HRR as a new method to investigate ns-motions in DNA, providing a general approach to study ns-motions in nucleic acids at atomic resolution. We will discover the fundamental motions in DNA, how they are affected by SSBs and lead to recognition by PARP-1.

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Ile-de-France Ile-de-France Paris
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