Determination of Membrane Protein Structure by Rotational Resonance NMR: Bacteriorhodopsin

Abstract

Rotationally resonant magnetization exchange, a new nuclear magnetic resonance (NMR) technique for measuring internuclear distances between like spins in solids, was used to determine the distance between the C-8 and C-18 carbons of retinal in two model compounds and in the membrane protein bacteriorhodopsin. Magnetization transfer between inequivalent spins with an isotropic shift separation, delta, is driven by magic angle spinning at a speed omega r that matches the rotational resonance condition delta = n omega r, where n is a small integer. The distances measured in this way for both the 6-s-cis- and 6-s-trans-retinoic acid model compounds agreed well with crystallographically known distances. In bacteriorhodopsin the exchange trajectory between C-8 and C-18 was in good agreement with the internuclear distance for a 6-s-trans configuration [4.2 angstroms (A)] and inconsistent with that for a 6-s-cis configuration (3.1 A). The results illustrate that rotational resonance can be used for structural studies in membrane proteins and in other situations where diffraction and solution NMR techniques yield limited information.

A First-Principles Study of 11-Cis-Retinal: Modelling the Chromophore-Protein Interaction In Rhodopsin

“…Here we discuss the experimental results obtained by the NMR methods. Verdegem et al have determined the nuclear distances C10-C20 (r 10,20 ) and C11-C20 (r 11,20 ) in the chromophore by using rotational resonance MAS NMR, which allows the measurement of internuclear distances between spin 1/2 nuclei (Creuzet, McDermott, Gebhard, van der Hoef et al, 1987;Verdegem, Helmle, Lugtenburg and de Groot, 1997) They have obtained r 10,20 = 0.304 ± 0.15Å and r 11,20 = 2.93 ± 0.15Å, respectively. The r 10,20 and r 11,20 distance measurements provide direct evidence that a considerable out-of-plane distortion is present in the central region of the rhodopsin chromophore.…”

Dipolar truncation in magic-angle spinning NMR recoupling experiments

“…In spite of the advances in pulse sequence design, the successful application of recoupling methods to the accurate measurement of 13 C- 13 C distance in macromolecules has typically been possible only when employing either specifically labeled spin-pair samples [26][27][28][29][30][31][32][33][34][35][36][37] or frequency-selective recoupling methods, [38][39][40][41][42][43][44][45][46][47] experiments in which a single internuclear distance is measured at a time. Although highly accurate ͑Ϯ5% is not difficult to achieve͒, both of these approaches are of limited practical utility in protein structure determinations that benefit from the availability of hundreds of constraints even for small proteins.…”

Using Sodium Cation Organization To Study the Phase Behavior of Bicelle Solutions

“…One particularly important approach involves using molecular constructs such as aqueous phase biologically friendly liquid crystal solvents such as the magnetically orientable Pf1 bacteriophage, 1 bacteriorhodopsin purple membranes, 2,3 or the phospholipid bicelles [4][5][6] to interrupt the isotropic tumbling of solutes and hence reintroduce chemical structure dependent features to the NMR spectrum of the solute. Of these media, the phospholipid bicelles are attractive due to their surface charge and ability to behave as cell membrane mimics.…”