NMR spectroscopic and analytical ultracentrifuge analysis of membrane protein detergent complexes

Maslennikov, Innokentiy; Kefala, Georgia; Johnson, Casey; Riek, Roland; Choe, Senyon; Kwiatkowski, Witek

doi:10.1186/1472-6807-7-74

Cited by 28 publications

(32 citation statements)

References 29 publications

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“…They have the same head group as phospholipids, but a single hydrophobic tail, which leads to the formation of micelles rather than bilayers. This is the main reason why these detergents are extremely efficient in E. coli membrane solubilization, 14,15 and foscholine-12 (FC12) with 12 methylene hydrophobic groups is one of the most effective membrane solubilization agents. FC12 has also been extremely useful and popular in NMR studies of membrane proteins aiming to obtain highresolution spectra in mixed micelles, 16 and has been shown to play a crucial role in refolding misfolded membrane proteins.…”

Section: Introductionmentioning

confidence: 99%

Structures of the OmpF porin crystallized in the presence of foscholine‐12

et al. 2010

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The endogenous Escherichia coli porin OmpF was crystallized as an accidental byproduct of our efforts to express, purify, and crystallize the E. coli integral membrane protein KdpD in the presence of foscholine-12 (FC12). FC12 is widely used in membrane protein studies, but no crystal structure of a protein that was both purified and crystallized with this detergent has been reported in the Protein Data Bank. Crystallization screening for KdpD yielded two different crystals of contaminating protein OmpF. Here, we report two OmpF structures, the first membrane protein crystal structures for which extraction, purification, and crystallization were done exclusively with FC12. The first structure was refined in space group P21 with cell parameters a 5 136.7 Å , b 5 210.5 Å , c 5 137 Å , and b 5 100.5°, and the resolution of 3.8 Å . The second structure was solved at the resolution of 4.4 Å and was refined in the P321 space group, with unit cell parameters a 5 215.5 Å , b 5 215.5 Å , c 5 137.5 Å , and c 5 120°. Both crystal forms show novel crystal packing, in which the building block is a tetrahedral arrangement of four trimers. Additionally, we discuss the use of FC12 for membrane protein crystallization and structure determination, as well as the problem of the OmpF contamination for membrane proteins overexpressed in E. coli.

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Section: Introductionmentioning

confidence: 99%

Structures of the OmpF porin crystallized in the presence of foscholine‐12

et al. 2010

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“…In use of different detergents, membrane proteins tend to exhibit varying functional activity, homogeneity, and stability. Inappropriate detergent exchange or detergentto-protein ratio usually results in inconsistent sample preparation and uncertainty of the protein -detergent complex structural homogeneity [26]. Thus, the use of a proper detergent is important for the efficient solubilization of membrane proteins for extraction and separation.…”

Section: Solubilization Of Membrane Proteinsmentioning

confidence: 99%

Microseparation of membrane proteins

Zhang¹,

Feng²,

Du³

et al. 2009

J of Separation Science

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Membrane proteins are generally of natural low abundance and insoluble to aqueous solutions. Thus, investigation of membrane proteins is relatively hampered since efficient separation of membrane proteins are rather challenging. Microseparation approaches certainly play an essential role in fulfilling such demanding investigations due to their significant advantages of high throughput, reduced separation time, and low sample consumption. In this review, recent developments of microseparation are documented with aspects of three key separation methods, including CE, micro-LC and microchip. Preparation of membrane proteins prior to separation is also discussed, including solubilization and preconcentration. Certainly, more applications of microseparation approaches in membrane protein separations can be expected in the near future.

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“…A detergent that is effective for extraction of an intrinsic protein from its membrane frequently turns out not to be ideal for subsequent characterization; in these cases a detergent exchange procedure is employed after the initial isolation. 14 Thus, for example, Triton X-100 is often effective for disruption of a biological membrane and concomitant “capture” of embedded proteins in a native-like state, but this detergent has generally not been amenable to crystallization. 15 Therefore, a sample solubilized with Triton X-100 can have the detergent component exchanged for more crystallization-prone examples such as DDM, OG or LDAO, which may not have been capable extracting the protein from its native membrane.…”

Section: Introductionmentioning

confidence: 99%

Crystallographic Characterization of N-Oxide Tripod Amphiphiles

2010

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Tripod amphiphiles are designed to promote the solubilization and stabilization of intrinsic membrane proteins in aqueous solution; facilitation of crystallization is a long-range goal. Membrane proteins are subjects of extensive interest because of their critical biological roles, but proteins of this type can be difficult to study because of their low solubility in water. The nonionic detergents that are typically used to achieve solubility can have the unintended effect of causing protein denaturation. Tripod amphiphiles differ from conventional detergents in that the lipophilic segment contains a branchpoint, and previous work has shown that this unusual amphiphilic architecture can be advantageous relative to traditional detergent structures. Here we report the crystal structures of several tripod amphiphiles that contain an N-oxide hydrophilic group. The data suggest that tripods can adapt themselves to a nonpolar surface by altering the hydrophobic appendage that projects toward that surface and their overall orientation relative to that surface. Although it is not possible to draw firm conclusions regarding amphiphile association in solution from crystallographic data, trends observed among the packing patterns reported here suggest design strategies to be implemented in future studies.

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NMR spectroscopic and analytical ultracentrifuge analysis of membrane protein detergent complexes

Cited by 28 publications

References 29 publications

Structures of the OmpF porin crystallized in the presence of foscholine‐12

Structures of the OmpF porin crystallized in the presence of foscholine‐12

Microseparation of membrane proteins

Crystallographic Characterization of N-Oxide Tripod Amphiphiles

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