Hydration State of Single Cytochrome c Monolayers on Soft Interfaces via Neutron Interferometry

Kneller, Larry R.; Edwards, Ann; Nordgren, C. Erik; Blasie, J. Kent; Berk, N. F.; Krueger, Susan; Majkrzak, C. F.

doi:10.1016/s0006-3495(01)76197-8

Cited by 19 publications

(24 citation statements)

References 19 publications

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“…X-ray and neutron reflectivity, enhanced by interferometry [15–17], can be utilized to characterize the profile structures of the chemisorbed bio-organic overlayers formed on the surface of inorganic multilayer substrates at each stage of their fabrication by either the DA or SA approach [18,19]. The “profile structure” is a projection of the overlayer’s three-dimensional (3D) structure parallel to the plane of the membrane onto the normal to that plane, thereby averaging over the in-plane structure of the overlayer.…”

Section: Introductionmentioning

confidence: 99%

Profile structures of the voltage-sensor domain and the voltage-gated K+-channel vectorially oriented in a single phospholipid bilayer membrane at the solid-vapor and solid-liquid interfaces determined by x-ray interferometry

et al. 2011

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One subunit of the prokaryotic voltage-gated potassium ion channel from Aeropyrum pernix (KvAP) is comprised of six transmembrane α helices, of which S1–S4 form the voltage-sensor domain (VSD) and S5 and S6 contribute to the pore domain (PD) of the functional homotetramer. However, the mechanism of electromechanical coupling interconverting the closed-to-open (i.e., nonconducting-to-K+-conducting) states remains undetermined. Here, we have vectorially oriented the detergent (OG)-solubilized VSD in single monolayers by two independent approaches, namely “directed-assembly” and “self-assembly,” to achieve a high in-plane density. Both utilize Ni coordination chemistry to tether the protein to an alkylated inorganic surface via its C-terminal His6 tag. Subsequently, the detergent is replaced by phospholipid (POPC) via exchange, intended to reconstitute a phospholipid bilayer environment for the protein. X-ray interferometry, in which interference with a multilayer reference structure is used to both enhance and phase the specular x-ray reflectivity from the tethered single membrane, was used to determine directly the electron density profile structures of the VSD protein solvated by detergent versus phospholipid, and with either a moist He (moderate hydration) or bulk aqueous buffer (high hydration) environment to preserve a native structure conformation. Difference electron density profiles, with respect to the multilayer substrate itself, for the VSD-OG monolayer and VSD-POPC membranes at both the solid-vapor and solid-liquid interfaces, reveal the profile structures of the VSD protein dominating these profiles and further indicate a successful reconstitution of a lipid bilayer environment. The self-assembly approach was similarly extended to the intact full-length KvAP channel for comparison. The spatial extent and asymmetry in the profile structures of both proteins confirm their unidirectional vectorial orientation within the reconstituted membrane and indicate retention of the protein’s folded three-dimensional tertiary structure upon completion of membrane bilayer reconstitution. Moreover, the resulting high in-plane density of vectorially oriented protein within a fully hydrated single phospholipid bilayer membrane at the solid-liquid interface will enable investigation of their conformational states as a function of the transmembrane electric potential.

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Section: Introductionmentioning

confidence: 99%

Profile structures of the voltage-sensor domain and the voltage-gated K+-channel vectorially oriented in a single phospholipid bilayer membrane at the solid-vapor and solid-liquid interfaces determined by x-ray interferometry

et al. 2011

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“…In the absence of a solid, recently developed, model-independent refinement methods may be used to derive these profiles with no a priori assumptions for monolayers at liquid-vapor or liquid-liquid interfaces [10]. When a solid is present, well-developed interferometric methods may be used [11]. We may apply these techniques to neutron reflectivity data collected from two monolayers prepared identically except that one contains an unlabeled (all 1 H) peptide while the other contains a peptide labeled with a single perdeuterated (i.e., maximally 1 H → 2 H substituted) residue at a pre-selected position in its amino acid sequence.…”

Section: Introductionmentioning

confidence: 99%

Specular neutron reflectivity and the structure of artificial protein maquettes vectorially oriented at interfaces

et al. 2004

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Artificial peptides can be designed to possess a variety of functionalities. If these peptides can be ordered in an ensemble, the functionality can impart macroscopic material properties to the ensemble. Neutron reflectivity is shown to be an effective probe of the intramolecular structures of such peptides vectorially oriented at an interface, key to ensuring that the designed molecular structures translate into the desired material properties of the interface. A model-independent method is utilized to analyze the neutron reflectivity from an alkylated, di- alpha -helical peptide, containing perdeuterated leucine residues at one or two pre-selected positions, in mixed Langmuir monolayers with a phospholipid. The results presented here are more definitive than prior work employing x-ray reflectivity. They show explicitly that the di-helical peptide retains its designed alpha -helical secondary structure at the interface, when oriented perpendicular to the interface at high surface pressure, with the helices projecting into the aqueous subphase without penetrating the layer of phospholipid headgroups.

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“…[31±33] the effect of temperature on the adsorption of hen egg white lysozyme at the silica ± water interface, [34] the hydration of cytochrome C tethered to Fe/Si or Fe/Au/Si surfaces, [35] and the sequential adsorption of fibrinogen and albumin. [9] One problem of neutron reflectivity in biology is the small contrast between the adsorbed molecule and the solvent.…”

Section: Neutron Reflectivitymentioning

confidence: 99%

Computer Simulations and Neutron Reflectivity of Proteins at Interfaces

Mungikar

Forciniti

2002

ChemPhysChem

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Hydration State of Single Cytochrome c Monolayers on Soft Interfaces via Neutron Interferometry

Cited by 19 publications

References 19 publications

Profile structures of the voltage-sensor domain and the voltage-gated K+-channel vectorially oriented in a single phospholipid bilayer membrane at the solid-vapor and solid-liquid interfaces determined by x-ray interferometry

Profile structures of the voltage-sensor domain and the voltage-gated K+-channel vectorially oriented in a single phospholipid bilayer membrane at the solid-vapor and solid-liquid interfaces determined by x-ray interferometry

Specular neutron reflectivity and the structure of artificial protein maquettes vectorially oriented at interfaces

Computer Simulations and Neutron Reflectivity of Proteins at Interfaces

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