Overview

Our laboratory is broadly focused on Nucleic Acid Nanotechnology and involves the design, construction, and application of Nucleic Acid-based nanoparticles. This is a vigorous and rapidly emerging field with strong potential for application in diverse disciplines ranging from nanoelectronics to nanomedicine. Our particular attention is devoted to RNA molecules, which not only encode genetic information but they also actively participate in various intracellular functions. This includes gene expression regulation through sophisticated mechanisms, thereby expanding RNA’s traditional role as a genetic messenger and revealing it as a functionally versatile molecule. Small interfering and microRNAs (siRNAs and miRNAs, respectively), ribozymes, rib switches, ribosomal RNA, transport RNA (tRNA) are only a few examples of non-coding RNA (ncRNA) elements that play diverse roles in mediating gene expression. To achieve their functions, RNAs fold into complex three dimensional (3D) architectures. Inspired by these natural 3D RNA elements, the development of artificial Nucleic Acid complexes with fine-tuned physicochemical properties is our main research interest.

Projects

Our research projects are highly interdisciplinary combining chemistry, biology, physics and material science. Our ultimate goal is the development of new strategies for controlled self-assembly of functional Nucleic Acid nanoparticles with implications in areas as diverse as nanoelectronics, biosensing, and nanomedicine. Projects provide students with extensive training in Nucleic Acid 3D design and various biochemical and phisycochemical techniques including DNA/RNA chemical labeling, Polymerase Chain Reaction (PRC), RNA and 2’F-modified RNA synthesis, ElectroMobility Shift Assays (EMSA), binding assays, UV-melting assays, Enzymatic resistance assays, protein over-expression and isolation. The following instruments are routinely exploited:  Fluorescence and UV- Vis absorption spectrophotometers, Dynamic Light Scattering (DLS), Surface Plasmon Resonance (SPR), Atomic Force Microscopy (AFM), Transmission Electron Microscopy (TEM).

Nanobiotechnology
In this project, we implement three basic RNA nanotechnology approaches to construct functional and robust RNA nanoparticles. (i) Computational approach includes in silico (computer-aided) design of RNA nanoobjects (2D and 3D geometries) using naturally occurring RNA building blocks. (ii) Experimental approach includes RNA synthesis, self-assembly, and structural characterization of the designed RNA nanoparticles. Lastly (iii), depending on the choice of application, these nanoparticles can be functionalized with various of biologically active compounds, used as a nanocontainers, utilized in construction of patterned arrays etc.

RNA computing
​The creation of the biomolecule-based computer is one of the most challenging yet fascinating tasks in synthetic biology and has intrigued great research attention. The major components of a computing device include an arithmetic logic unit, the control unit, memory, and inputs/outputs. The driving core of the fully operational computer system lies at the basis of Boolean logic gates and, therefore, to create a biocomputer one has to first generate biomolecular logic gates. Fluorogenic RNA and DNA aptamers are attractive candidates to fulfill this task due to their various light emitting properties and structural diversity. In our lab we implement secondary structure prediction algorithms to design and construct RNA and RNA/DNA complexes that are able to perform basic AND, OR, NOR, NAND logistic operations. Our goal is to achieve a nucleic acid based combinatorial logic system with unbiased emission output from a set of short oligonucleotide inputs.

Thermodynamics of RNA non-canonical interactions
Structures of large RNA molecules, such as the ribosomal RNAs (rRNA), have revealed that no more than ~70% of all basepairs in structured RNAs are cis Watson-Crick (cWW) AU, GC, or GU pairs. The rest are non-WC basepairs and together with base-stacking and base-backbone they serve to locally structure the nominally single-stranded hairpin, internal, and multi-helix junction “loops” evident in 2D structures to form modular 3D motifs, many of which are recurrent. These interactions expose functional groups along the base edges, especially the base Watson-Crick and Sugar edges, that mediate long-range RNA interactions crucial for RNA folding. The “loops” also form specific binding sites for proteins, small molecule ligands, metals, and other RNAs.
The basepair isostericity hypothesis is fundamental for understanding the rules of sequence adaptation during RNA evolution. Isostericity indicates which basepairs can potentially substitute for each other while preserving the 3D structure of a motif, but says nothing about the thermodynamic implications of the substitution. Only a limited information is available about the relative effects of isosteric vs. non-isosteric base substitutions on the thermodynamic stabilities of well-structured RNA 3D motifs, or whether particular base combinations are favored, depending on the optimal growth temperature to which the organism is adapted. This project addresses these fundamental questions.