Photophysical and structural investigation of a (Py)A-modified adenine cluster: its potential use for fluorescent DNA probes exhibiting distinct emission color changes.

In this study, we found a (Py)A-modified adenine cluster (A-cluster), a minimum fluorescent unit for significant emission wavelength changes, and investigated its photophysical and structural properties. The basic A-cluster unit was an adenine-pentad duplex containing stacked (Py)A pairs in the center aligned in an antiparallel manner. Spectral analysis of the A-cluster revealed remarkable reddish fluorescence with a large Stokes shift (∼195 nm) and a long life-time constant (31 ns), originated from exciton states formed by (Py)A pairs and neighboring adenines. Structurally, the exciton state of the A-cluster exhibited unusually high stability, relative to that of other five-mismatched duplexes, as a result of stabilization through strong stacking interactions (zipper-like structure) of the mismatched A-A and (Py)A pairs, rather than through traditional Watson-Crick base pairing. These spectral and structural properties of the A-clusters were specific to the adenine bases and highly disturbed by introducing other bases (T, G, and especially C) or an abasic site into the A-cluster, whereas they were enhanced through synergistic effects in systems containing multiple A-clusters. As a minimum unit for these unique properties, finally, the A-cluster was exploited as a fluorescent probing system for specific nucleic acid sequences, such as miR-21, accompanying distinct fluorescence color changes from blue to red. These findings indicated the potential utility of the A-cluster as a part of fluorescent probes exhibiting clear signaling upon micro-environmental changes.