Biochemistry and Biophysics

Listed below are the research projects that our researchers are involved in.

RNA methyltransferases

We are interested in understanding the molecular basis underlying the protein-RNA interactions involved in a broad range of cellular activities. One of our recent efforts concerns the structure-function of RNA modifying enzymes. Through a structural biology approach, combined with biochemical and cellular assays, we aim to elucidate the molecular mechanisms of these enzymes at atomic resolution. For instance, our structure-function characterization of the ZCCHC4 protein, a 28S rRNA-specific m6A methyltransferase, reveals an autoinhibitory mechanism underpinning the specific recognition between ZCCHC4 and the RNA substrate with a stem-loop structure. Our long-term goal is to elucidate how individual RNA modifying enzymes have evolved into distinct substrate specificity.

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Translation, RNA-binding proteins, RNA disease

Controlled protein translation is an integral component of regulated gene expression, and allows cells to respond rapidly to changing conditions. Translation is heavily regulated during its initiation, where a plethora of protein factors coordinate with ribosomal subunits and initiator tRNA to ensure efficient and faithful identification of the mRNA start codon. We aim to decipher the molecular and biophysical mechanisms of this highly intricate, exquisitely dynamic process. We focus on how mRNA–factor interactions guide recognition of the mRNA 5ʹ cap by initiation factor eIF4F, how mRNA is loaded onto the 40S subunit, and how the resulting 48S pre-initiation complex scans the mRNA leader to locate the start codon. We probe these molecular mechanisms with a combination of biochemical and single-molecule biophysics approaches. We aim to understand how the distinct properties of different mRNAs modulate initiation, how disease states reconfigure initiation dynamics, and how therapeutic interventions mitigate dysfunction.


Transcription-Translation Coupling, Structure Of Nascent RNA, Gene Expression

The absence of a nuclear membrane in bacteria allows the RNA to be translated while being transcribed. The occurrence of both processes on the same RNA at the same time results in a dynamic interplay between RNA polymerase and ribosome, coupling transcription and translation. This coupling determines the structure of the nascent RNA between the polymerase and the ribosome. RNA structures can affect transcription and translation. The signal for intrinsic termination of transcription consists of a stem loop followed by six-eight Us. Shifting of the reading frame of translation is promoted by pseudo loops within the mRNA. The effect of RNA structures on transcription and translation is well documented but is less understood when both processes are coupled. To elucidate this interplay, we are bringing to bear single molecule and cryo-EM technology, in collaboration with Seán O’Leary, UCR, and Richard Ebright, Rutgers University, respectively.


Circulating RNA, RNA structure, RNA modification

Non-coding RNAs participate in gene expression regulation, and are highly relevant to pathological development. They are stably present in diverse body fluids and can be sampled non-invasively for clinical tests. Thus, circulating RNAs have great potential to be disease biomarkers. To improve discovery of effective RNA markers, one of our research focuses is to detect circulating RNAs, in particular, the RNAs enclosed in extracellular vesicles (EVs). EVs are important cell-cell communicators that can transport RNAs to recipient cells to carry out specific functions. Analyzing EV RNAs could reveal effective circulating RNA markers for disease diagnosis and prognosis. We are also interested in analysis of nucleic acid folding, another factor affecting gene expression. Working with our collaborator, we have developed synthetic receptor arrays for differentiation of diverse DNA folding motifs like G-quadruplex, hairpins, i-motifs, and triplexes. We are working on expanding this method to analyze RNA structures.


Nanopore-based RNA Sensing and Biophysics

The translocation of molecules, particularly protein and RNA, are ubiquitous and vital to cellular life.  Furthermore, protein and mRNA undergo a plethora of modifications which diversifies their function and creates sub-populations that are difficult to detect using bulk ensemble measurements.  Using a single nanopore as a sensor, RNA molecules can be probed and characterized by size, shape, and charge.  Based on the physiochemical shift in molecular properties which is typical of binding events (e.g. complex formation), folding (e.g. secondary structure), and mutations (e.g. stability of secondary structure), a host of single molecule biophysics can be probed and provide unique insight into RNA biology. Our lab focuses on technology/technique development which can push the limits of single molecule sensing and improve on the time resolution of sensing (~microsecond resolution), capture rate (i.e. sensitivity), and specificity of measuring essential biophysical properties of molecules.