Determination of the number of detergent molecules associated with the reaction center protein isolated from the photosynthetic bacterium <i>Rhodopseudomonas viridis</i>

Gast, P.; Hemelrijk, Petra W.; Hoff, A.J.

doi:10.1016/0014-5793(94)80625-x

Cited by 42 publications

(23 citation statements)

References 17 publications

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“…After minimization, the protein complex was embedded into a micelle that was composed of LDAO detergent molecules. Following the estimates of Gast et al 33 , 260 detergent molecules were used to solvate the TM portion of the protein. After insertion, around 187 Na + and 188 Cl − ions were added to the water phase to create a 0.2 M salt solution.…”

Section: Methodsmentioning

confidence: 99%

Visualizing a protein quake with time-resolved X-ray scattering at a free-electron laser

et al. 2014

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A ‘protein quake’ describes the hypothesis that proteins rapidly dissipate energy through quake like structural motions. Here we measure ultrafast structural changes in the Blastochloris viridis photosynthetic reaction center following multi-photon excitation using time-resolved wide angle X-ray scattering at an X-ray free electron laser. A global conformational change arises within picoseconds, which precedes the propagation of heat through the protein. This motion is damped within a hundred picoseconds.

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Section: Methodsmentioning

confidence: 99%

Visualizing a protein quake with time-resolved X-ray scattering at a free-electron laser

et al. 2014

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“…RC from R. sphaeroidees R26 were isolated as described in [ll]; they contained more than 90% QH acceptor. LDAO detergent was removed according to a method described earlier [9]. Briefly, the amount of detergent necessary to resolubilize a certain amount of detergent-free RC was determined by measuring the light scattering at 500 nm.…”

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confidence: 99%

The Association of Different Detergents with the Photosynthetic Reaction Center Protein of Rhodobacter sphaeroides R26 and the Effects on its Photochemistry

Gast

Hemelrijk

Gorkom

et al. 1996

European Journal of Biochemistry

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Detergent-free reaction centers from Rhodobacter sphaeroides R26 were used to study the solubilization of reaction centers in various detergents and their effects on reaction center photochemistry. 500Z 100 n-octyl-P-D-glucopyranoside or 51 i 5 Triton X-100 molecules were associated with one reaction center. For N,N-alkylamine N-oxide detergents with chain lengths in the range from 8-13 carbon atoms, the number of detergent molecules associated with the reaction centers increased with decreasing chain length. The amount of detergent molecules associated with the reaction centers decreased almost tenfold if the pH was increased from pH 6 to pH 10. The addition of 5 % 1,2,3-heptanetriol to various detergents lowered the detergentheaction center ratio by a factor of two compared to that for the pure detergent. The detergent concentration at which solubilization of the reaction center occurs was close to the critical micelle concentration for all detergents studied. The absorption band at 865 nm of the primary donor in the reaction center shifts to 846nm when detergent was removed from the reaction center; upon resolubilization with various detergents, this band shifts back to 865 nm. In 80-90% of the detergent-free reaction centers, the secondary electron transfer from QA to QB was inhibited; this electron transfer was restored after re-addition of detergent.Keywords: photosynthesis ; membrane protein ; detergent: crystallization; Rhodobacter sphaeroides.Crystallization of detergent-solubilized membrane proteins is complicated by the randomly ordered detergent molecules surrounding the proteins. The success of crystallization of membrane proteins depends on optimizing hydrophobic and hydrophilic interactions between the protein molecules. The type of detergent plays an important role in this process. The length of the detergent alkyl chain determines the thickness of the detergent belt around the protein. This belt should be as thin as possible to increase the number of contact sites between the membrane proteins ; however, the detergent belt should not be too small, so that the detergents can form bridges between the proteins inside the crystal structure [I]. Additionally, the polar head group of the detergent should be small to ensure optimal hydrophilic interactions between the protein molecules [l].The first successful crystallization of a membrane protein was achieved with the photosynthetic reaction center (RC) protein from the purple bacterium Rhodopseudomonas viridis [2]. The addition of 1-3% of the amphiphillic molecule 1,2,3-heptanetriol was essential for the formation of crystals. This led Michel [3] to postulate that the dimensions of the micellar disk surrounding the protein are fine-tuned by 1,2,3-heptanetriol so as to fit into the crystal structure and to increase proteidprotein interactions. Measurements on the micellar radius of N,Ndimethyl dodecylamine N-oxide (DodNOMe, or LDAO) in solution have shown a decrease of the radius upon addition of 1,2,3-heptanetriol The size of this detergent belt is d...

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“…20, 30, 40 10. 20, 30 (20) 4.5 Roth et al (1989) studied the size of the detergent ring surrounding this reaction centre in its crystallized form and the LDAO/reaction centre (RC) ratio value was found to be 110, a value more than two times lower than the value of 260 needed for the RC's solubilization (Gast et al, 1994). In work carried out on the structure of the detergent phase in the Rhodohacter sphaeroides (strain Y) reaction centre crystals (Roth et al, 1991) it was reported that /3-octylglucoside formed a beltshaped micelle around the hydrophobic 'waist' of the complex with interconnections between belts via micellar bridges.…”

Section: Introductionmentioning

confidence: 99%

A novel approach for the crystallization of soluble proteins using non-ionic surfactants

Mustafa

Derrick

Tiddy

et al. 1998

Acta Crystallogr D Biol Cryst

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Crystallization trials using three polyoxyethylene surfactants as precipitating agents are described. Of the eight soluble proteins screened, five were successfully crystallized at the first attempt. These included lysozyme, catalase, ferritin, ribonuclease A and ubiquitin. Further work suggested that these surfactants could also be suitable for cryo-crystallographic analysis of crystals. At the concentrations used in the crystallization trials [10-40%(v/v)], they are capable of promoting the formation of non-crystalline glasses at cryogenic temperatures (77 K). This would facilitate crystal mounting and allow the minimization of crystal irradiation damage. Results from this study also suggest that proteins remain stable at high concentrations of these surfactants [40%(w/v)] and over long time periods (>1 month). A number of membrane proteins were also screened for crystallization. These included photosystems I and II and light harvesting complexes I and II from spinach and bacteriorhodopsin from Halobacterium halobium. The trials were unsuccessful both in the absence and presence of heptane-l,2,3-triol and over a wide range of surfactant concentrations.

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Determination of the number of detergent molecules associated with the reaction center protein isolated from the photosynthetic bacterium Rhodopseudomonas viridis

Cited by 42 publications

References 17 publications

Visualizing a protein quake with time-resolved X-ray scattering at a free-electron laser

Visualizing a protein quake with time-resolved X-ray scattering at a free-electron laser

The Association of Different Detergents with the Photosynthetic Reaction Center Protein of Rhodobacter sphaeroides R26 and the Effects on its Photochemistry

A novel approach for the crystallization of soluble proteins using non-ionic surfactants

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