A summary of my ALK2 project goals.

I thought I’d spend a blog post just talking about the various paths of my research and try and tie it all together. A lot of my experiments do dot around these various arms of investigation and so one experimental post doesn’t necessarily tie in directly with the previous one or the next one and Read More …

Optimising seeding of normal and FOP cells for a later assay – prettier than expected

Further down the line I want to be able to test the activation of the BMP signalling pathway downstream of the ALK2 receptor. For this I plan to use the alkaline phosphatase gene as a readout – it’s a well known gene activated downstream of BMP signalling so a lot of easy to use assays Read More …

A new ALK2 structure, and proof that mistakes in the lab aren’t always a bad thing

Hello everyone and happy New Year! I really, really wanted to end the year on a high note in December before I went off on holiday, but as it turned out, I didn’t have enough time to set up 18 plates and write a blog post on the last day. So I’ll just have to Read More …

Determining the nanoBRET ALK2 IC50 values of 24 new ACVR1/ALK2 inhibitors

In an effort to develop clinical compounds for Diffused Intrinsic Pontine Glioma (DIPG) treatment, new analogues of ACVR1/ALK2 inhibitors are continuously synthesised by Ontario Institute for Cancer Research (OICR) and Charles River Laboratories (CRL). I will provide prompt feedback of the cellular assay results to guide their design of new compounds. I determined the potency Read More …

Purification and crystallisation of an ALK2/ACVR2/FKPB12 complex.

One problem with working with the isolated kinase domains of ALK2 and it’s type II binding partners, is that the affinity between the kinase domains alone is insufficient to allow complex formation. We know that the two proteins associate with each other in vivo and are co-localised by the domains on the outside of the Read More …

Screening ACVR1 inhibitors on mutant and non-mutant ACVR1 DIPG cells – effectiveness may vary

Hi there! The last month of my life was taken over by making sure my PhD first year report was beautifully polished, but I have returned with results of a small compound screen: These are all compounds that Jong Fu has already tested with his assays so we know they effectively inhibit ACVR1, but I Read More …

Update on crystal hits, XChem and more purification

Things have been really busy in the lab the last few weeks. I’ve done a couple of purifications of ACVR1 (ALK2), which I’ll post in Zenodo (getting loads of protein now), but probably won’t write up the details of every purification every single time anymore, as it’s the same process every time for this construct. Read More …

Generating mutations in the GS domain of ALK2

This week one of the things I’ve been working on is the final set of mutations in the GS domain of Alk2 to help us unpick the influence of phosphorylation on function. That, however, is a bit of a mouthful so let’s unpack it a bit. I said in my last blog post that we Read More …

Some structure pictures for your viewing pleasure

So I reported here a couple of weeks ago that I’d obtained datasets for ACVR1 bound to M4K2117 and M4K2121, at 1.25 and 2 Å resolution, respectively, and I put up a link to the data. I’m now refining the data and got some pretty good structures so far. These are what the current stats look Read More …

Phosphorylation of SMAD1 – rates as a function of Alk2 mutations and type II association.

One of the questions to be asked when looking at the mutation in ALK2 that causes FOP, is how exactly does that mutation cause the disease. ALK2 we know is part of the BMP signalling pathway responsible for bone formation and the mutation results in excess bone formation – thus we can summarise that somehow Read More …