SICOR: Subgraph Isomorphism Comparison of RNA Secondary Structures.

RNA aptamer selection during SELEX experiments builds on secondary structural diversity. Advanced structural comparison methods can focus this diversity. We develop SICOR, which uses probabilistic subgraph isomorphisms for graph distances between RNA secondary structure graphs. SICOR outperforms other comparison methods and is applicable to many structural comparisons in experimental design.

A light-responsive RNA aptamer for an azobenzene derivative.

Regulation of complex biological networks has proven to be a key bottleneck in synthetic biology. Interactions between the structurally flexible RNA and various other molecules in the form of riboswitches have shown a high-regulation specificity and efficiency and synthetic riboswitches have filled the toolbox of devices in many synthetic biology applications. Here we report the development of a novel, small molecule binding RNA aptamer, whose binding is dependent on light-induced change of conformation of its small molecule ligand. As ligand we chose an azobenzene because of its reliable photoswitchability and modified it with chloramphenicol for a better interaction with RNA. The synthesis of the ligand 'azoCm' was followed by extensive biophysical analysis regarding its stability and photoswitchability. RNA aptamers were identified after several cycles of in vitro selection and then studied regarding their binding specificity and affinity toward the ligand. We show the successful development of an RNA aptamer that selectively binds to only the trans photoisomer of azoCm with a KD of 545 nM. As the aptamer cannot bind to the irradiated ligand (λ = 365 nm), a light-selective RNA binding system is provided. Further studies may now result in the engineering of a reliable, light-responsible riboswitch.

Small-Molecule-Binding Riboswitches.

RNA is a versatile biomolecule capable of transferring information, taking on distinct three-dimensional shapes, and reacting to ambient conditions. RNA molecules utilize a wide range of mechanisms to control gene expression. An example of such regulation is riboswitches. Consisting exclusively of RNA, they are able to control important metabolic processes, thus providing an elegant and efficient RNA-only regulation system. Existing across all domains of life, riboswitches appear to represent one of the most highly conserved mechanisms for the regulation of a broad range of biochemical pathways. Through binding of a wide range of small-molecule ligands to their so-called aptamer domain, riboswitches undergo a conformational change in their downstream "expression platform." In consequence, the pattern of gene expression changes, which in turn results in increased or decreased protein production. Riboswitches unite the sensing and transduction of a signal that can directly be coupled to the metabolism of the cell; thus they constitute a very potent regulatory mechanism for many organisms. Highly specific RNA-binding domains not only occur in vivo but can also be evolved by means of the SELEX (systematic evolution of ligands by exponential enrichment) method, which allows in vitro selection of aptamers against almost any ligand. Coupling of these aptamers with an expression platform has led to the development of synthetic riboswitches, a highly active research field of great relevance and immense potential. The aim of this review is to summarize developments in the riboswitch field over the last decade and address key questions of recent research.