Druggability and Genetic Variability of the Catalytic and Allosteric Sites of SARS-CoV-2 nsp14 Across Coronaviruses and SARS-CoV-2 Samples

Hello and happy new year! I am back from my winter break and starting to get back to research. If you have followed my previous posts, you know that I have posted the druggability and genetic variability analysis of several protein targets of SARS-CoV-2. There are a few targets that I will be posting in Read More …

Project Overview: Introduction to WDR12 and WDR55 Proteins

I am currently completing my fourth-year thesis in Dr. Dalia Barsyte-Lovejoy’s lab. I have a strong interest in both pharmacology and biochemistry. I am interested in learning about biochemical pathways in the body, with the goal of finding compounds that can target those pathways and for them to potentially develop into therapeutics. My project focuses Read More …

Distinguishing between catalytic and non-catalytic pockets in the ligandable human genome

My goal is to find druggable pockets in human enzymes that are non-catalytic. These non-catalytic druggable pockets may then be exploited for proximity pharmacology (ProxPharm), a novel paradigm in drug discovery where chimeric compounds bring two proteins in close proximity to elicit an effect of one protein on the other1. For example, PROTACs simultaneously bind Read More …

Druggability and Genetic Variability of the ATPase Site and Central Channel of SARS-CoV-2 nsp13 Helicase Across Coronaviruses and SARS-CoV-2 Samples – Post 23

One of the important enzymes in the replication cycle of the SARS-CoV-2 virus is the helicase, which is also known as non-structural protein 13 (nsp13). During the viral life cycle, the holo-RNA-dependent RNA polymerase, also known as nsp12, is thought to coordinate with several additional factors, including the nsp13 helicase (Snijder et al., 2016; Sola Read More …

Druggability and Genetic Variability of the ADP-bound Pocket of SARS-CoV-2 RNA-dependent RNA polymerase NiRAN domain Across Coronaviruses and SARS-CoV-2 Samples – Post 22

The RNA-dependent RNA-polymerase (RdRp) also known as non-structural protein 12 (nsp12) is the target of antiviral agent remdesivir. Nsp12 has an important role in viral genome replication and transcription. (Chen et al., 2020)  Chen et al. identified a new pocket on the N-terminal extension of nsp12 occupied by ADP-Mg2+ after solving the structure of the helicase-polymerase complex. This ADP-bound pocket is on the N-terminal nidovirus RdRp-associated nucleotidyltransferase (NiRAN) domain which Read More …

Interrogating PR/SET domains for the ability to bind SAH

This is an excerpt from my PhD thesis where I investigated 13 human PR/SET domains from the PRDM protein family and assessed their ability to bind to a fluorinated SAH analog. The research helps to identify which of the PRDMs could be methyltransferases. This work was carried out in the laboratory of Dr. Cheryl Arrowsmith Read More …

Production and purification of Ectodomain of SARS-CoV-2 Spike (S) protein

Production and purification of Ectodomain of SARS-CoV-2 Spike (S) protein Two different expression plasmids for prefusion S ectodomain residues 1−1208 of 2019-nCoV S (GenBank: MN908947) with proline substitutions (2P) at residues 986 and 987(1) and with six proline substitutions (HexaPro (at residues 817, 892, 899, 942, 986 and 987 (2), a “GSAS” substitution at the Read More …

Development and Optimization of an IQF USP5 Activity Assay

In a previous post, I described a ubiquitin-rhodamine110 (UbRho110) assay, and showed that a ZnF-UBD ligand antagonizes USP5 catalytic activity. UbRho110 is not a native substrate, and validation with native polyubiquitin species in an orthogonal assay is required. To address this, I developed and optimized an internally quenched fluorophore pair (IQF) assay. You can see Read More …

Genetic Variability at An Allosteric site of SARS-CoV-2 RNA-dependent RNA polymerase Across Coronaviruses and SARS-CoV-2 Samples – Post 21

SARS-CoV-2 is a positive-strand RNA virus, depending on its multi-subunit machinery to replicate its RNA. This machinery is known as RNA-dependent RNA polymerase (RdRp). The catalytic subunit of RdRp, which is the core component of this machinery, is called nsp12. Nsp12 has little catalytic activity on its own and relies on accessory subunits to have Read More …

FBXW11 interaction with β-catenin – cellular assay

Background The SCF (SKP1-CUL1-F-box protein) E3 ubiquitin ligase is a complex of proteins that helps facilitate the ubiquitination of substrates for targeted proteasomal degradation. FBXW11 is F-box containing protein that has 11 WD-repeats and is a substrate recognition component of the SCF complex. FBXW11 recognizes and binds its substrates, including beta-catenin, at phosphorylated sites or Read More …