There's a new toy in the lab! I’ve been dreaming of acquiring a Prometheus for a long time (since my PhD), and we finally managed to purchase one at the #LaJollaInstitute. I can't wait to start characterizing our glycoproteins with it! #biophysics #cryoem #proteinfolding - ThreadSky

dzyla.bsky.social • 103 days ago

There's a new toy in the lab! I’ve been dreaming of acquiring a Prometheus for a long time (since my PhD), and we finally managed to purchase one at the #LaJollaInstitute. I can't wait to start characterizing our glycoproteins with it! #biophysics #cryoem #proteinfolding

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gerg77.bsky.social•102 days ago

Absolutely love those instruments! Not sure if you'll be doing any membrane protein work, but check out this app note if you're interested: https://cube-biotech.com/media/8c/9e/cf/1725626988/Appnote-2024_Cube%20Biotech-Nanotemper.pdf

dzyla.bsky.social•102 days ago

That's a great idea! We do a lot of proteins currently and buffer optimization is one of the tasks which we will use Prometheus for 😄

automnenine.bsky.social•102 days ago

How effective is buffer optimization for non-membrane proteins? I've always wanted this to work, but I've never had any luck for what we do.

gerg77.bsky.social•102 days ago

Tough question! My unsatisfying answer is you'd just have to try on a per protein basis.

Off the top of my head, this could be a fun workflow:
1. prep protein in a generic OK buffer, get it to ~1 mg/mL
2. dilute 1:1 in a pre-formulated buffer screen (make it by hand in 96-well)
3. Check on a panta

dzyla.bsky.social•102 days ago

Before Prometheus, I used standard DSF testing pH & salt. Surprisingly, pH had no effect, but high salt did. Switching to 800 mM NaCl buffer raised protein melting temp by almost 10°C. Definitely worth trying!

gerg77.bsky.social•102 days ago

Probably worth revisiting, on the off chance that the cypro dye was muting pH-dependent changes

dzyla.bsky.social•102 days ago

Will do! We have recorded some data using nanoDSF, and the results were very complimentary, so I am not worried about it too much. Anyway, a great era of protein condition optimization is ahead of us!

osvalb.bsky.social•99 days ago

We've developed some tools to analyse dsf data. Please check them out and don't doubt to contact us 😁 https://spc.embl-hamburg.de/

dzyla.bsky.social•99 days ago

That is cool! I will spread the word about it in the lab

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