Conformation of Membrane Proteins: Bacteriorhodopsin
Abstract
One of the main goals of this research project was accomplished. A water soluble derivative of bacteriorhodopsin (BR) was synthesized. An activated species of methoxypolyethylene glycol (MeOPEG) was synthesized, yielding 2-0- methoxypolyethylene glycol-N-hydroxy succinimyl carbonate. (MeO-PEG-SC). MeOPEG- SC was coupled with the purple membrane (PM) of Halobacterium hilobium to yield MeO-PEG-PM. This product was centrifuged and purified by washing with H20 (76 % conversion), dissolved in buffer with 5 % sodium dodecylsulphate (SDS), (1: 1) and PAGE Electrophoresis performed. The separated MEO-PEG-BR band was recovered from the gel by electroelution, and the solution lyophilized. This material was dissolved in H20 and the SDS removed by passing the solution through an Extra- gel column (Pierce) and eluted with P04 buffer pH 7.0. The final solution was centrifuged at 200,000 g, yielding a clear water-soluble solution of MeO-PEGBR. The product could be reconstituted into miscells which were capable of proton pumping. The MeO-PEG-BR reconstituted into vesicles had physical-chemical properties identical with the original PM. The circular dichroism spectra, ultraviolet and visible spectrum, and fluorescence spectrum were identical to that of the PM. Proteins, Membranes, Conformation, Bacteriorhodopsin.
Document Details
- Document Type
- Technical Report
- Publication Date
- Dec 09, 1991
- Accession Number
- ADA248455
Entities
People
- Gerald D. Fasman
Organizations
- Brandeis University