Hello everyone,
We need to do large scale yeast protein preps and so far I have not
found a convenient way of doing it. Glass beads on large scale is
painful, not effective (only ~ 50% breakage) and expensive.
Spheroplasting is long, poorly reproducible and leads to
more than usual proteolysis. Liquid nitorogen method in our hands
is messy as hell (half of the yeasts end up all over the place) and
not very effective either. French press is convenient but even less
effective than glass beads.
Pierce offers what's supposed to be a great achiever that overcomes
almost all the problems above. "Y-PER" reagent. Some super-duper
secret recipie which you use to resuspend cells, incubate for 20 min
at rt, and after that ~ 90% cytoplasmic proteins is said to be in the
supernatant. Obviously, some kind of detergent. We need to test
protein for activity after expression, and it is not at all impossible
that the unknown Y-PER will simply kill the protein (Pierce offers
bGal as an example of the enzyme but this is not serious - bGal
is one of the sturdiest enzymes around). I also don't feel like spending
$150 to find out whether any of Pierce's calims are true.
So the question is - what is it? Known things (that to me only tell it is
a fairly concentrated solution of some long chain fatty acid-containing
detergent) :
- _is_ detergent;
- "may start to solidify at 17.9C";
- Specific gravity ~ as water (1.03);
- better soluble in methanol than in water; solubility in cold water is
limited;
- insoluble in diethyl ether and n-octanol;
- stable at rt;
- reactive with acids and somewhat reactive with oxidizing agents.
As usual, it is probably something trivial. Even then, I would even
buy the thing if I knew in advance what it is all about.
Any ideas?
- Dima
