non-denaturing gels on thylakoid pigment-protein complexes

Thomas Thomas at hastingsmac2.fas.harvard.edu
Tue Sep 24 10:59:06 EST 1996


In article <526gm7$3ku at net.bio.net>, leslie at ibg.uit.no (leslie wanner) wrote:

> I am interested in a simple non-denaturing gel system for separating
> thylakoid membrane pigment protein complexes.  I have read much about the
> use of Deriphat-PAGE gels, which sound good, but where does one buy or
> acquire Deriphat?  Alternatively, are there other similar detergents that
> are easier to come by aAnd give similar results?  I am interested in names
> and sources of detergents, and in protocols or references to protocols.
>
> Thanks for the help,
>
> Leslie Wanner
> University of Tromso
> Norway
> e-mail: leslie at ibg.uit.no
> FAX: +47 776 45600

Look at Dunahay and Staehlin, Plant Physiol 78: 606-613 (1985) and 80:
429-434 (1986), also Peter and Thornber, I think the ref is JBC 266:
16745-54 (1991).
Detergents of the maltoside- (e.g. Decylmaltoside) and glucoside- type
(e.g. octylglucoside) are very mild detergents used successfully in the
electrophoresis of PS-complexes (often in combination). Try variation of
the detergent to protein ratio and the detergent 1/detergent 2 ratio.
Preparation of thylakoid membranes should be carried out on ice, gel
electrophoresis should be done at 4oG, if possible (just put your chamber
in the fridge and close the door - sound awful, but works; set up the
power supply right next to the fridge-outside, of course.). Preparation of
the thylakoid membranes (should be starch free) is crucial as well as the
solubilization step, include a potease inhibitor like Pefabloc.

Thomas

--

Sender:  Thomas Urbig
         Harvard University
         Institute for Molecular and Cellular Biology
         16 Divinity Ave.
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         Email:  Thomas at hastingslab.harvard.edu





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