Question
Asked 23rd Sep, 2014

Does anyone have experience with isolation of purple membrane from halobacterium?

Is there any specific mutated strain regarding this isolation?

All Answers (2)

Ravi Kant Upadhyay
Deen Dayal Upadhyay Gorakhpur University
Isolation of the cell membrane of Halobacterium halobium and its fractionation into red and purple membrane by a via aqueous-two-phase system.Thepurple membrane contains a chromoprotein with a retinal prosthetic group similar to rhodopsin, the chromprotein found in the visual systems of higher invertebrates and vertebrates. The described purple membrane isolation procedures yield a highly purified preparation as determined by transmitting electron microscopy and gel electrophoresis. Critical analysis of the absorption spectra of the purple membrane is also employed to establish criteria of purity for the preparation. The visible absorption spectra of the purified purple membrane preparation in buffer is  used to a maximum at 559 nm which shifted to 567 nm on light exposure. 
James Coker
University of Maryland Global Campus
You can do the separation via a sucrose density gradient centrifugation.  It is not technically difficult to do but generating the sucrose gradient may take a bit of time.  You will probably need a gradient of about 20-45% (w/v).  The purple membrane should probably form a band around 25% in this set up.  If you use the right tubes you can puncture the side with a needle/syringe and extract the purple membrane directly.
About a mutant strain to use, you should look into using the Halobacterium S9 strain (http://www.pnas.org/content/98/5/2521.full).  It constitutively produces purple membrane (as opposed to getting turned on after the lowering of oxygen).

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