Exploring Nucleobase Modifications in Oligonucleotide Analogues for Use as Environmentally Responsive Fluorophores and Beyond

Abstract

Over the past two decades, it has become abundantly clear that nucleic acid biochemistry, especially with respect to RNA, is more convoluted and complex than previously appreciated. Indeed, the application and exploitation of nucleic acids beyond their predestined role as the medium for storage and transmission of genetic information to the treatment and study of diseases has been achieved. In other areas of endeavor, utilization of nucleic acids as a probe molecule requires that they possess a reporter group. The reporter group of choice is often a luminophore because fluorescence spectroscopy has emerged as an indispensable tool to probe the structural and functional properties of modified nucleic acids. The scope of this review spans research done in the Hudson lab at The University of Western Ontario and is focused on modified pyrimidine nucleobases and their applications as environmentally sensitive fluorophores, base discriminating fluorophores, and in service of antisense applications as well as tantalizing new results as G‐quadruplex destabilizing agents. While this review is a focused personal account, particularly influential work of colleagues in the chemistry community will be highlighted. The intention is not to make a comprehensive review, citations to the existing excellent reviews are given, any omission of the wonderful and impactful work being done by others globally is not intentional. Thus, this review will briefly introduce the context of our work, summarize what has been accomplished and finish with the prospects of future developments.

Constructing Artificial Nucleobase Compilation to Enable Precise Molecular Medicine†

“…The phenyl‐substituted pC analogues, such as 6‐phenylpyrrolocytidine (PhpC) and boPhpC, are structurally conservative and highly fluorescent cytidine analogues, which are the most commonly used probes for fluorescence‐based assays. [ 92 ] Wahba et al . reported that PhpC‐containing RNA can act as a molecular beacon to report HIV‐1 RT Ribonuclease H activity and has the potential to screen the inhibitors of HIV‐RT RNase H. [ 93 ] The same group also reported that PhpC‐containing small interfering RNAs (siRNAs) can be used to investigate cellular distribution and mechanism of siRNAs.…”

Screening for Novel Fluorescent Nucleobase Analogues Using Computational and Experimental Methods: 2-Amino-6-chloro-8-vinylpurine (2A6Cl8VP) as a Case Study

“…In order to probe the structure and dynamics of DNA and RNA sequences and their interaction with other biological molecules, structural nucleobase analogues have been developed with high-emission quantum yields. − These structural analogues are called fluorescent nucleic acid base analogues (FBAs). A summary of recent developments in the field of FBAs can be found in these articles. − Two of the oldest known fluorescent base analogues are 2-aminopurine (2AP) and ethenoadenine (εA), both of which are fluorescent analogues of adenine. 8-Vinyladenine is a relatively new adenine analogue introduced by Mely et al Fluorescent analogues are also known for G, , C, − and U. , Besides their use in understanding biological nucleic acid properties, they have also been used in understanding and making synthetic DNA-based devices, such as photoinduced electron transfer through DNA nanowires. − New improved FBAs are needed with better single photon and multiphoton emission properties − to facilitate single molecule microscopy studies in vitro and in living cells.…”

Chimeric GFP–uracil based molecular rotor fluorophores

“…Thus, intrinsic nucleobase modifications are done to impart fluorescence properties to the nucleobase while preserving their canonical base-pairing abilities. 4 Surprisingly, it has been found that large number of such intrinsic modifications to the nucleobase gives fluorophores with emissions in the blue region of the visible spectrum 5 and/or are not sufficiently bright for use in real-time applications. For applications in biological systems, it is desirable to have (i) emissions of fluorophores bathochromically shifted from the blue region of the visible spectrum, (ii) practically usable brightness and (iii) environmental sensitivity to changes in the microenvironment.…”