Solid-state NMR investigations of the bacteriorhodopsin photocycle

Bacteriorhodopsin (bR) is a heptahelical integral membrane protein with a retinylidene prosthetic group from H. salinarum. The protein creates a proton gradient across the cell membrane by harvesting light energy. Proton transfer events can be segregated into transfers between the Schiff base of the chromophore and the cytoplasmic and extracellular half-channels. Critical to pump function is the switch in accessibility between the two half-channels. Models of the accessibility switch invoke protein or chromophore structural changes. This thesis describes the application of solid-state Nuclear Magnetic Resonance (NMR) to investigate each of the competing models in the native purple membrane environment. Conformational changes of the chromophore have been characterized by correlation of 13C-1H and 5N-1H dipolar interactions to determine chromophore dihedral angles in multiple photocycle intermediates. The resting state of the protein is found to contain a pre-loaded chromophore, with distortion about the Schiff base linkage and the neighboring C15-C14 bond. Changes of the dihedral about the C15-C14 were characterized during the photocycle, although the timing of the changes precludes their direct involvement in the accessibility switch. Putative protein conformation changes at the X-Pro peptide bonds were probed via the peptide 1 N and 13C chemical shifts. Proposed roles of the X-Pro bonds discriminate between structural and dynamic roles during the photocycle.

Department dc:contributor.department

Massachusetts Institute of Technology. Dept. of Chemistry.

Grantor dc:publisher

Massachusetts Institute of Technology

Year dc:date.issued

2002

Author dc:creator

• Lansing, Jonathan C. (Jonathan Clifford), 1974-

Advisor dc:contributor.advisor

• Robert G. Grifffin.

• Chemistry.

Statement dc:rights

• M.I.T. theses are protected by copyright. They may be viewed from this source for any purpose, but reproduction or distribution in any format is prohibited without written permission. See provided URL for inquiries about permission.

Licence dc:rights.uri

• http://dspace.mit.edu/handle/1721.1/7582

Language dc:language.iso

eng

Handle dc:identifier.uri

http://hdl.handle.net/1721.1/8050

OAI identifier oai:identifier

oai:dspace.mit.edu:1721.1/8050