IUBio

mysterious lysozyme or what?

P.C. PC at no.email.sorry
Wed Jan 28 11:28:17 EST 2004


Kaj, which pH did you use for the Q and SP sepharoses (for lysozyme)?

lysozyme has several disulfides; depending on Cys reduction/oxydation 
status and pH, the protein might bind to anion echange and elute in 
several peaks. With all its Cys ingaged in disulfides the pI will be 
above 10!!

best,

Peter

kaj.stenberg at helsinki.fi.invalid wrote:
> P.C. <PC at no.email.sorry> wrote:
> 
>>I find it very strange that lysozyme would bind at all to Q sepharose 
>>(not that I tried it myself). Can it be that the real peak actually did 
>>not elute from SP yet?
> 
> 
> I redid the experiment. From Q-sepharose 2-3 overlapping peaks in
> gradient. From SP one peak in gradient. SDS-PAGE shows identical bands
> in all relevant fractions, corresponding to lysozyme size.
> 
> 
>>Are your columns fresh and clean? If not, wash them in 8M GuHCl.
>>Or NaOH as APB recommends.
> 
> 
> They were freshly packed, hence the lysozyme test.
> 
> 
>>And finally, are you sure that the vial that you used for chromatography 
>>was actually chicken egg lysozyme and not chicken egg albumin? (please 
>>do not take offence, it is just another thing to check). 
> 
> 
> No offence taken, strange things happen. I checked the vial, it is
> lysozyme (hen egg white, 3 x crystallized). 
> 
> I only wanted to use the Q-sepharose column and it worked in my
> isolation so I leave the mysterious binding to SP in the folder 
> "mysteries". But if someone has any explanation to offer I would 
> be grateful.
> 




More information about the Proteins mailing list

Send comments to us at biosci-help [At] net.bio.net