Protein from Yeast

[Bernard P. Murray, PhD]

In article <3703A2E6.64A0 at po-box.mcgill.ca>, maarts at po-box.mcgill.ca wrote:

> I don't know if anyone can help but we are looking for a good protocol
> to extract protein from yeast.  Currently we are sonicating the yeast
> with glass beads and only getting 100-200micrograms per 50mL of
> culture.  Not very efficient.  Any suggestions would be greatly
> appreciated. 
> thanks
> Michelle

For small amounts of culture for soluble protein (eg. luciferase)
I extract with zirconia/silica beads.  For larger volumes and
for subcellular fractionation I make sphaeroplasts with lytic
enzyme and then sonicate in hypotonic buffer.  Whatever method
you use don't forget to include plenty of protease inhibitors.
The yeast methods in Current Protocols give a reasonable
starting methodology but you then need to adapt it for your
specific purpose.
     Do you mean 100 - 200 micro-g of total protein or of
your protein of interest?
          Bernard
-- 
Bernard P. Murray, PhD
Dept. Cell. Mol. Pharmacol., UCSF, San Francisco, USA