Measurement of Proton/M412 Ratios in Suspensions of Purple and White Membrane from Halobacterium Halobium
We have studied isolated purple membrane from Halobacterium halobium strain S9 and white membrane from the mutant strain JW-5, making careful measurements of the proton to M412 stoichiometry of our preparations, and also examining them by Quasi Elastic Light Scattering (QELS) and by negative staining electron microscopy (EM), to assess extent of aggregation and hydrodynamic radius.
In the past decade there have been varying reports of the stoichiometry of proton release from purple membrane in suspension and protons pumped after its incorporation into phospholipid vesicles. Interpretation of the results, especially when by a group other than that which carried out the experiments, has sometimes been clouded by a misunderstanding of the differences between the various measurements involved and also by a lack of recognition that the aggregation state of membranes will affect the measured ratios.
In an attempt to resolve these misunderstandings, we have investigated the H+/M412 ratio, measured in suspensions of purple membranes by the laser flash induced response of pH indicator dyes. We have found this ratio to reflect the aggregation state of the membrane, assessed on the same preparations by QELS (and EM, see accompanying chapter: Lefort-Tran et al.). We have also found our measurements of proton release stoichiometry on retinal reconstituted white membrane suspensions to give much higher H+/M412 ratios than the same measurements on purple membrane suspensions.
KeywordsTrypsin Treatment Purple Membrane Proton Release Bulk Aqueous Phase Quasi Elastic Light Scattering
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