Infrared spectroscopic study of photoreceptor membrane and purple membrane. Protein secondary structure and hydrogen deuterium exchange.

Downer, Nancy W.; Bruchman, T J; Hazzard, Jo H.

doi:10.1016/s0021-9258(17)35695-8

Cited by 80 publications

(26 citation statements)

References 31 publications

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“…2), suggest that not all of them are easily accessible. This is in agreement with previous infrared spectroscopic investigations where even at full hydration 71% of the BR amide groups remain inexchangeable (Downer et al, 1986).…”

Section: Discussionsupporting

confidence: 93%

Deuterium Solid-State NMR Investigations of Exchange Labeled Oriented Purple Membranes at Different Hydration Levels

Bechinger

Weik

2003

Biophysical Journal

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Oriented purple membranes were equilibrated under controlled (2)H(2)O relative humidity ranging from 15% to 93% and introduced into the magnetic field of an NMR spectrometer with the membrane normal parallel to the magnetic field direction. Deuterium solid-state NMR spectra of these samples resolved four deuteron populations. Deuterons that have exchanged with amide protons of the protein exhibited a broad spectral line shape (<150 kHz). Furthermore, a broadened signal of deuterons tightly associated with protein and lipid is detected at low hydration, as well as two additional water populations that were present when the samples were equilibrated at >/=75% relative humidity. These latter ones are characterized by narrow quadrupolar splittings (<2.5 kHz) and orientation-dependent chemical shifts. Their deuterium relaxation times, measured as a function of temperature, indicate correlation times in the fast regime (10(-10) s) and activation energies of 13 kJ/mol (at 86% relative humidity). Differences in T(1) and T(2) relaxation together with small residual quadrupole splittings show that the mobility of the deuterons is anisotropic. The occurrence of these mobile water populations at high levels of purple membrane hydration (>/=75% relative humidity) correlate with proton pumping activity of bacteriorhodopsin, the fast kinetics of M-decay in the bacteriorhodopsin photocycle, and structural alterations of the protein during the M-state, which have been described previously.

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Section: Discussionsupporting

confidence: 93%

Deuterium Solid-State NMR Investigations of Exchange Labeled Oriented Purple Membranes at Different Hydration Levels

Bechinger

Weik

2003

Biophysical Journal

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“…In this process, membranes are suspended in buffered D 2 O and rehydrated in a controlled D 2 O atmosphere exposing amide protons from residues in loops at the surface of the protein to the aqueous environment. These amide protons are able to exchange with deuterons, whereas those that are buried in the membrane or, involved in secondary structure hydrogen bonding, remain protonated (Downer et al, 1986). Met-68 is shown in a high resolution electron diffraction structure of the surface of bacteriorhodopsin as being part of a b-sheet in the BC loop (Kimura et al, 1997); however, it is not involved in hydrogen bonding and infrared analysis of bacteriorhodopsin secondary structure has suggested the presence of disturbed b-structure (Downer et al, 1986).…”

Section: Mas (600)mentioning

confidence: 99%

“…These amide protons are able to exchange with deuterons, whereas those that are buried in the membrane or, involved in secondary structure hydrogen bonding, remain protonated (Downer et al, 1986). Met-68 is shown in a high resolution electron diffraction structure of the surface of bacteriorhodopsin as being part of a b-sheet in the BC loop (Kimura et al, 1997); however, it is not involved in hydrogen bonding and infrared analysis of bacteriorhodopsin secondary structure has suggested the presence of disturbed b-structure (Downer et al, 1986). It is likely, therefore, that the amide proton of Met-68 can be exchanged in the presence of D 2 O.…”

Section: Mas (600)mentioning

confidence: 99%

Identifying Anisotropic Constraints in Multiply Labeled Bacteriorhodopsin by 15N MAOSS NMR: A General Approach to Structural Studies of Membrane Proteins

Mason

Grage

Straus

et al. 2004

Biophysical Journal

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Structural models of membrane proteins can be refined with sets of multiple orientation constraints derived from structural NMR studies of specifically labeled amino acids. The magic angle oriented sample spinning (MAOSS) NMR approach was used to determine a set of orientational constraints in bacteriorhodopsin (bR) in the purple membrane (PM). This method combines the benefits of magic angle spinning (MAS), i.e., improved sensitivity and resolution, with the ability to measure the orientation of anisotropic interactions, which provide important structural information. The nine methionine residues in bacteriorhodopsin were isotopically (15)N labeled and spectra simplified by deuterium exchange before cross-polarization magic angle spinning (CPMAS) experiments. The orientation of the principal axes of the (15)N chemical shift anisotropy (CSA) tensors was determined with respect to the membrane normal for five of six residual resonances by analysis of relative spinning sideband intensities. The applicability of this approach to large proteins embedded in a membrane environment is discussed in light of these results.

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“…Much less is known about structural changes occurring in the peptide backbone of bR. BR consists of a peripheral region where H/D exchange of backbone amide peptide groups occurs rapidly and a core region where the backbone amide peptide groups are largely inaccessible to H/D exchange (18,19). Polarized FTIR spectroscopy combined with H/D exchange has shown that this core region consists primarily of R-helical structure oriented predominantly perpendicular to the membrane plane and buried largely within the lipid bilayer (19,20).…”

mentioning

confidence: 99%

Conformational Changes in the Core Structure of Bacteriorhodopsin

et al. 1998

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Bacteriorhodopsin (bR) is the light-driven proton pump found in the purple membrane of Halobacterium salinarium. In this work, structural changes occurring during the bR photocycle in the core structure of bR, which is normally inaccessible to hydrogen/deuterium (H/D) exchange, have been probed. FTIR difference bands due to vibrations of peptide groups in the core region of bR have been assigned by reconstituting and regenerating delipidated bR in the presence of D2O. Exposure of bR to D2O even after long periods causes only a partial shift of the amide II band due to peptide NH --> ND exchange only of peripheral peptide structure. However, the amide II band completely downshifts when reconstitution/regeneration of bR is performed in the presence of D2O, indicating that almost the entire core backbone structure of bR undergoes H/D exchange. Peripheral regions can then be reexchanged in H2O, leaving the core backbone region deuterated. Low-temperature FTIR difference spectra on these core-deuterated samples reveal that peptide groups in the core region respond to retinal isomerization as early as the K intermediate. By formation of the M intermediate, infrared differences in the amide I region are dominated by much larger structural changes occurring in the core structure. In the amide II region, difference bands appear upon K formation and increase upon M formation which are similar to those observed upon the cooling of bacteriorhodopsin. This work shows that retinal isomerization induces conformational changes in the bacteriorhodopsin core structure during the early photocycle which may involve an increase in the strength of intramolecular alpha-helical hydrogen bonds.

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Infrared spectroscopic study of photoreceptor membrane and purple membrane. Protein secondary structure and hydrogen deuterium exchange.

Cited by 80 publications

References 31 publications

Deuterium Solid-State NMR Investigations of Exchange Labeled Oriented Purple Membranes at Different Hydration Levels

Deuterium Solid-State NMR Investigations of Exchange Labeled Oriented Purple Membranes at Different Hydration Levels

Identifying Anisotropic Constraints in Multiply Labeled Bacteriorhodopsin by 15N MAOSS NMR: A General Approach to Structural Studies of Membrane Proteins

Conformational Changes in the Core Structure of Bacteriorhodopsin

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