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231225s2018 xx |||||o 00| ||eng c |
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|a 10.1021/acs.langmuir.8b01503
|2 doi
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|a pubmed24n0962.xml
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|a (NLM)30239200
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|a DE-627
|b ger
|c DE-627
|e rakwb
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|a eng
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| 100 |
1 |
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|a Skar-Gislinge, Nicholas
|e verfasserin
|4 aut
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| 245 |
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|a Comprehensive Study of the Self-Assembly of Phospholipid Nanodiscs
|b What Determines Their Shape and Stoichiometry?
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|c 2018
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|a Text
|b txt
|2 rdacontent
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|a ƒaComputermedien
|b c
|2 rdamedia
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|a ƒa Online-Ressource
|b cr
|2 rdacarrier
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| 500 |
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|a Date Completed 19.12.2018
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|a Date Revised 19.12.2018
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|a published: Print-Electronic
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|a Citation Status PubMed-not-MEDLINE
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|a Phospholipid nanodiscs have quickly become a widely used platform for studies of membrane proteins. However, the molecular self-assembly process that ultimately should place a membrane protein inside a nanodisc is not well understood. This poses a challenge for a successful high-yield reconstitution of general membrane proteins into nanodiscs. In the present work, the self-assembly process of POPC-MSP1D1 nanodiscs was carefully investigated by systematically modulating the reconstitution parameters and probing the effect with a small-angle X-ray scattering analysis of the resulting nanodiscs. First, it was established that nanodiscs prepared using the standard protocol followed a narrow but significant size distribution and that the formed nanodiscs were stable at room temperature over a time range of about a week. Systematic variation of the POPC/MSP1D1 stoichiometry of the reconstitution mixture showed that a ratio of less than 75:1 resulted in lipid-poor nanodiscs, whereas ratios of 75:1 and larger resulted in nanodiscs with constant POPC/MSP1D1 ratios of 60:1. A central step in the self-assembly process consists in adding detergent-absorbing resin beads to the reconstitution mixture to remove the reconstitution detergent. Surprisingly, it was found that this step did not play a significant role for the shape and stoichiometry of the formed nanodiscs. Finally, the effect of the choice of detergent used in the reconstitution process was investigated. It was found that detergent type is a central determining factor for the shape and stoichiometry of the formed nanodiscs. A significantly increasing POPC/MSP1D1 stoichiometry of the formed nanodiscs was observed as the reconstitution detergent type is changed in the order: Tween80, DDM, Triton X-100, OG, CHAPS, Tween20, and Cholate, but with no simple correlation to the characteristics of the detergent. This emphasizes that the detergents optimal for solution storage and crystallization of membrane proteins, in particular DDM, should not be used alone for nanodisc reconstitution. However, our data also show that when applying mixtures of the reconstitution detergent cholate and the storage detergents DDM or OG, cholate dominates the reconstitution process and nanodiscs are obtained, which resemble those formed without storage detergents
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|a Journal Article
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|a Research Support, Non-U.S. Gov't
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|a Johansen, Nicolai Tidemand
|e verfasserin
|4 aut
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1 |
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|a Høiberg-Nielsen, Rasmus
|e verfasserin
|4 aut
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| 700 |
1 |
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|a Arleth, Lise
|e verfasserin
|4 aut
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| 773 |
0 |
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|i Enthalten in
|t Langmuir : the ACS journal of surfaces and colloids
|d 1992
|g 34(2018), 42 vom: 23. Okt., Seite 12569-12582
|w (DE-627)NLM098181009
|x 1520-5827
|7 nnns
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| 773 |
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|g volume:34
|g year:2018
|g number:42
|g day:23
|g month:10
|g pages:12569-12582
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|u http://dx.doi.org/10.1021/acs.langmuir.8b01503
|3 Volltext
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|d 34
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|e 42
|b 23
|c 10
|h 12569-12582
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