Year: 2018
Manuscript finished! Design and characterisation of mutant and wild-type huntingtin proteins produced from a toolkit of scalable eukaryotic expression systems
Hi folks! This past month or so I have been busy getting a manuscript draft prepared. This article details the HTT constructs we have made and shared with the community, how they can be used to make pure huntingtin protein of different polyQ lengths and characterisation of the protein samples that we made. We think this Read More …
Huntington Society of Canada 2018 Conference in Kelowna
Hi everyone! This weekend I am in rainy Kelowna for the Huntington Society of Canada National Conference. I am excited to meet all the participants and hear from scientists, clinicians, patients and other folks who attend this important meeting. On Saturday, I will be giving a short presentation on free and legal ways to access Read More …
Biophysical investigation of HTT and HTT-HAP40 using DSLS and SAXS
Hi folks! Over the past month or so, I have been conducting biophysical characterisation experiments of the HTT and HTT-HAP40 samples I am able to purify in the lab. I have used DSLS which measures thermal stability by monitoring protein aggregation over a temperature gradient. In an ongoing collaborative effort with Sasha Lemak and Lixin Read More …
Searching far and wide for proteins that bind huntingtin
Hi folks! I have just returned from a much needed vacation and catch up with my family back in the UK. I am now back in the lab and catching up on everything that has been happening whilst I have been away. Before I left, I didn’t quite have time to update you all on Read More …
Expression and purification of the HTT-HAP40 complex with expanded polyQ (Q54)
Previously I showed that I could reproduce the expression and purification of HTT-HAP40, shown by Guo et al in their Nature paper earlier this year. Now I have been able to make this same complex but with polyQ-expanded HTT. It would seem that HAP40 also stabilises the Q54 HTT molecule as much as the Q23 HTT molecule Read More …
The ongoing quest for huntingtin interaction partners
This past week I have been replenishing my stocks of huntingtin samples making both the full-length Q42 protein as well as the full-length Q23 protein in complex with HAP40. The protocol for HTT-HAP40 complex generation is now very robust and the sample looks top notch. I have also tried to make HTT complexes with other Read More …
Starting new projects in huntingtin structural biology
Hi folks! This last month or so has been a bit of a whirlwind. I was ecstatic to receive the HDSA career development fellowship and we were also successful in our funding applications to the Huntington Society of Canada as well as the CHDI Foundation. There will be some slight redirection in the projects but Read More …
Full-length huntingtin protein aggregation and first attempts to co-purify HTT-HAP40 from insect cells
This past week, I have been completing experiments in two different threads: 1. I have used the different polyQ length huntingtin protein samples I made previously in an experiment which measures how the protein molecules aggregate (stick together) when heated. This technique is called differential static light scattering (DSLS). I saw no difference in the Read More …
New folks in open notebooking!
Aside from work in the lab, I have been busy promoting the cause of open notebooking at a number of different events around the world. This has been very rewarding and I have enjoyed engaging different audiences in the work our open notebook community is doing. I spoke recently at the Creative Commons Global Summit Read More …
Full-length and exon 1 huntingtin protein purification from bacteria, insect and mammalian cells
Recently I have been busy making lots of huntingtin protein samples for various experiments I have planned shortly to work out what the protein is doing with respect to binding DNA and other nucleic acid material. I have been using constructs we made in house which have either N-terminal or C-terminal FLAG tags and a Read More …