Isolation of Hydrophobic Membrane Proteins by Perfusion Chromatography—Purification of Photosystem II Reaction Centers from Spinach Chloroplasts
✍ Scribed by Margrit Roobol-Bóza; Bertil Andersson
- Publisher
- Elsevier Science
- Year
- 1996
- Tongue
- English
- Weight
- 129 KB
- Volume
- 235
- Category
- Article
- ISSN
- 0003-2697
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✦ Synopsis
Ion exchange perfusion chromatography has been teins requires the presence of detergent to solubilize introduced for the isolation of hydrophobic membrane the proteins from the lipid bilayer and to keep them protein complexes from thylakoid membranes of spinin solution during subsequent isolation. However, the ach chloroplasts. By using this chromatographic techpresence of detergent may result in damaging effects nique, previously shown to be useful for the rapid isoto the protein such as unfolding, loss of cofactors, inaclation of soluble proteins (Regnier, F. E. (1991) Nature tivation of catalytic functions, or modifications of spe-350, 634-635), we have been able to isolate oxygen cific protein-protein interactions. These problems beevolving photosystem II core complexes and photosyscome particularly apparent when detergent treatments tem II reaction center particles. Pure reaction centers are combined with time-consuming isolation procecould be isolated from photosystem II core complexes dures.
after a chromatographic step requiring only 6.5 min,
In this study we introduce the new technique of per-
which is a substantial improvement in comparisons fusion chromatography (2, 3) as a rapid procedure to with previous procedures. The entire preparation of isolate a hydrophobic membrane protein complex from photosystem II core complexes and reaction center photosynthetic membranes of plant chloroplasts. Perdesigned with two different types of pores to increase 1 This work has been supported by the Swedish Natural Science the mass transport while maintaining a very high reso-Research Council, the Go ¨ran Gustafsson Foundation for Natural Scilution and capacity (4). The large ''through pores'' ences and Medicine and the Jan Artur Ekstro ¨ms Foundation.