Discriminating Single Base Pair Mismatches in DNA Using Glutathione-Templated Copper Nanoclusters

A single nucleotide mismatch in a particular sequence of DNA is considered to play pivotal roles in various central biological processes and is associated with the development of several types of oncogene and genetic diseases. Hence, the identification of particular probes for DNA is of great interest in order to carry out cell imaging, drug delivery, and point of care diagnostic. Herein we report the binding interaction between DNA and tripeptide-functionalized luminescent copper nanoclusters (CuNCs) for recognition of single base pair mismatched (MM) double-stranded (ds) DNA from well-matched (WM) sequences. Isothermal titration calorimetry and UV-vis thermal denaturation established that substitution of a single well matched GC pair in 20 base pair (bp) DNA with single pyrimidine mismatches; viz. CA, CC, and CT mispair resulted in an enhanced binding affinity of CuNCs to ds DNA. Through fluorescence correlation spectroscopy, we estimated the precise values of the binding interaction at the single-molecule resolution. Analysis of autocorrelation curves, hydrodynamic radius, and association rate constants (K-ass) evidently established the higher affinity of CuNCs for the single pair MM dsDNA. The order of binding affinity of CuNCs is found to be CT MM > CA MM > CC MM > WM DNA. Competitive binding studies using Hoechst and ethidium bromide substantiate the groove binding mechanism.