Hi Lionel, A few musings/suggestions
If they are bound *inside* the protein, this suggests that the phospholipids might be very tightly bound. Do you have an affinity tag on your protein (e.g. His tag)? Perhaps you might immobilise the protein on a suitable resin and wash with copious amounts of detergent, then elute as you would normally? One can remove LPS from a prep this way using Triton X-114, perhaps this might work for you? I do agree that HIC might be useful as well. Failing that, and given the fact that these PLs are bound *in* perhaps the only way to completely remove them from a sample might be to denature the protein (Urea or Guanidine?), remove the lipids (HIC, Lipidex, immobilisation/washing as above) and then hope that the protein can be refolded without the PL present? Perhaps the PLs are an essential structural component? Hope this is helpful! Good luck! Dave Dr David C Briggs PhD http://about.me/david_briggs On 26 Jun 2014 17:16, "Lionel" <lionel.costen...@gmail.com> wrote: > Dear all, > > I would like to remove two phospholipids bound to (actually into) a > cytoplasmic protein. The protein was expressed in E. coli and the crystal > structure revealed the presence of two co-purified phospholipids (most > probably DPPE). > > A web search gave me three methods to remove bound lipids: > > - 1-butanol liquid–liquid extraction > - Lipidex 1000, VI column at 37°C > - HIC (aka Hydrophobic Interaction Chromatography) on a Phenyl HP column > in presence of 1M ammonium sulphate > > All are described in Velkov 2008 ( > http://www.sciencedirect.com/science/article/pii/S1570023208002390). > > I assume these delipidation methods would also work for phospholipid; and > I am more tempting by the last one, the milder one. > > I would appreciate any comment or practical advice. > > Best regards, > Lionel >
