Dear Andreas,
maybe the problem is rather rooted in your expression set up. If the
protein was soluble in the small scale test, it is not necessarily the
case in a fermenter run. The parameters may have changed so strongly that
your protein is now expressed mostly insoluble. To my experience,
Andreas, you probably know all this, but I only understood quite recently.
What happens is that as ice crystals form you get brine rejection, the
same thing that happens in the arctic when sea water freezes. Therefore
you can have protein concentrated in pockets of high salt. Fine for some
Dear all,
I've encountered people who refuse to freeze cells and always lyse fresh
pellets. Better protein, they say. I've never had reason to do so
myself, or even to believe in their voodoo. Up until now, maybe.
My protein expresses well and is almost all in the soluble fraction in
an
I have experience with some proteins that don't tolerate freeze-thawing
very well. It's hard to say exactly what the physical chemistry of this
is, but it probably relates to (1) aggregation due to high concentration
or protein or salts during the freezing process as water is removed,
and/or
