Pyrene–cyanine conjugates as multipurpose fluorescent probes for non-covalent recognition of ds-DNA, RNA and proteins

Abstract

Two novel conjugates of pyrene and cyanine were constructed by linking them with a rigid triazole–peptide linker. These new probes bind very strongly (with 0.1 μM affinity) to both ds-DNA(RNA) and proteins (BSA), giving significantly different fluorimetric responses: a strong pyrene emission change is highly selective for proteins and the “switch-on” of cyanine fluorescence is highly selective for DNA(RNA). Moreover, the new probes yield induced CD bands only with DNA/RNA, but not with BSA, which allowed an independent check of DNA presence in DNA/protein mixtures. Furthermore, these probes contain a FRET pair of chromophores, whereby FRET is silent in a free molecule solution and is activated by binding of the small molecule to the biomacromolecular target. The efficiency of FRET is to some extent related to the secondary structure of DNA/RNA and only for one of the probes is FRET activated in proteins. The two probes show distinctively different induced CD patterns in the 400–600 nm range (attributed to a different position of linker attachment on the cyanine core), allowing differentiation between various secondary structures of DNA or RNA, which are shown to be additionally enhanced by combining pyrene and cyanine into one molecule. Due to their low cytotoxicity and efficient cellular uptake, these probes are good candidates for further biological studies.