USP5 ZnF-UBD Co-Crystal Structure with Compound UBTR012574a

A Happy New Year to all! I am back from vacation and ready to jump back into my work with USP5. Great news- we solved the co-crystal structure of USP5 with a promising compound, UBTR012574a (Figure 1) (discussed in my last post), just before the Christmas holidays. You can check out details about co-crystal growth, Read More …

Expression and solubility test of SETDB1 catalytic domain constructs from insect (Sf9) cell expression systems (1)

In my previous post, I shared the expression and purification results of several constructs based on bacterial expression systems. Such experiments often fail as there are inherent complications with protein expression associated with bacterial expression systems. This was likely the case for most of our last batch samples. To tackle this problem, we decided to Read More …

UGGT: a fly-paper glycoprotein catching troublesome flies?

  About one quarter of the Eukaryotic genome encodes glycoproteins that are made in the Endoplasmic Reticulum (ER), and travel to special destinations such as the extracellular environment, the cell membrane or other organelles like the lysosomes. We can think of the ER as a nursery where baby glycoproteins quickly grow to attain their adult Read More …

Blog 2020_1: Scientific resolutions for 2020

A happy new year all, yes I know. It’s already mid-January and we are nearly all done with our resolutions. But what better way for me to start back to the open notebooking/blogging than to kick off with a top 10 list of my resolutions/hopes for 2020 (some science/some aspirational). Yes, I am back blogging Read More …

Initial cell potency screening of kinase STK17A/DRAK1 inhibitors

Protein STK17A or DRAK1 is a serine/threonine kinase that belongs to the death-associated protein kinase family (DAPK). This family is comprised of five members: DAPK1, DAPK2, DAPK3, DRAK1 and DRAK2, being the last two the least studied.  As part of our efforts to find inhibitors for understudied kinases, we recently disclosed a high quality chemical Read More …

Mass spec analysis of SMAD1 phosphorylation by ALK2 and ALK2 GS mutants in the presence of ACVR2.

I decided to test a set of my ALK2 GS loop phosphomutants for activity. As I’ve written about previously, I needed to optimise the concentrations of receptors and decided that 60uM was the right concentration to use as showing activity with the mutants and also being workable in the assay. I set up three different Read More …

Defining Epigenetic Drivers of Medulloblastoma – 03 – Dose Response of BAY-299

Update #03 – Performing Dose-response assays of BAY-299 on medulloblastoma   I have been screening multiple medulloblastoma cell lines including ONS-76, D425, D458, and Med8A with BAY-299 and its negative control probe.  So far, three out of four lines have demonstrated sensitivity to the BAY-299 probe consistent with the screening results (Defining Epigenetic Drivers of Read More …

Identifying mono methyl arginines with PTMScan® Mono-Methyl Arginine Motif [mme-RG] Kit from CST

We have used the PTMScan® Mono-Methyl Arginine Motif [mme-RG] Kit #12235 from CST in order to determine novel PRMT7 substrates. In preliminary experiments done few years ago by our pharma partner several novel potential PRMT7 substrates were discovered with the kit, including HSP70, which was further validated by different methods (https://www.biorxiv.org/content/10.1101/503136v2). In order to obtain biological replicates, Read More …

Purification of HTT N-HEAT_81-1643 and initial biophysical characterization

In the second purification round of N-HEAT_81-1643, I added an additional purification step with heparin resin. My aim with doing this additional step was to improve the purity of the HTT N-HEAT_81-1643 sample from nucleic acid material/other proteins. Similar experiments proved successful for full HTT+HAP40 sample (by Dr. Rachel Harding). Our results showed that the Read More …