The Role of the Detergent Micelle in Preserving the Structure of Membrane Proteins in the Gas Phase

Reading, Eamonn; Liko, Idlir; Allison, Timothy M.; Benesch, Justin L. P.; Laganowsky, Arthur; Robinson, Carol V.

doi:10.1002/anie.201411622

Cited by 126 publications

(173 citation statements)

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“…We therefore screened various detergents to identify one that requires lower activation energy to release the CcO complex intact from the detergent micelle. C 8 E 4 is considered a "harsher" detergent (i.e., its ability to substitute for natural lipids is low compared with DM or DDM) but it requires lower activation energy to release proteins from micelles in the gas phase (25).…”

Section: Ptms Are Located On the Peripheral Subunits And In The Dimericmentioning

confidence: 99%

Dimer interface of bovine cytochrome c oxidase is influenced by local posttranslational modifications and lipid binding

Liko

Degiacomi

Mohammed

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Bovine cytochrome c oxidase is an integral membrane protein complex comprising 13 protein subunits and associated lipids. Dimerization of the complex has been proposed; however, definitive evidence for the dimer is lacking. We used advanced mass spectrometry methods to investigate the oligomeric state of cytochrome c oxidase and the potential role of lipids and posttranslational modifications in its subunit interfaces. Mass spectrometry of the intact protein complex revealed that both the monomer and the dimer are stabilized by large lipid entities. We identified these lipid species from the purified protein complex, thus implying that they interact specifically with the enzyme. We further identified phosphorylation and acetylation sites of cytochrome c oxidase, located in the peripheral subunits and in the dimer interface, respectively. Comparing our phosphorylation and acetylation sites with those found in previous studies of bovine, mouse, rat, and human cytochrome c oxidase, we found that whereas some acetylation sites within the dimer interface are conserved, suggesting a role for regulation and stabilization of the dimer, phosphorylation sites were less conserved and more transient. Our results therefore provide insights into the locations and interactions of lipids with acetylated residues within the dimer interface of this enzyme, and thereby contribute to a better understanding of its structure in the natural membrane. Moreover dimeric cytochrome c oxidase, comprising 20 transmembrane, six extramembrane subunits, and associated lipids, represents the largest integral membrane protein complex that has been transferred via electrospray intact into the gas phase of a mass spectrometer, representing a significant technological advance.cytochrome c oxidase | mass spectrometry | lipids

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Section: Ptms Are Located On the Peripheral Subunits And In The Dimericmentioning

confidence: 99%

Dimer interface of bovine cytochrome c oxidase is influenced by local posttranslational modifications and lipid binding

Liko

Degiacomi

Mohammed

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

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“…Recently there has been an upsurge in the use of other non-ionic detergents for native MS analysis of MPs, for example, Triton X-100, tetraethylene glycol monooctyl ether (C8E4), octaethylene glycol monododecyl ether (C12E8), lauryldimethylamine N-oxide (LDAO), and n-octyl-b-D-glucoside (b-OG) (Fig. 4) as it has been shown that these detergent micelles dissociate at much lower activation energies, often resulting in MP ions that are more native-like [95][96][97]. These detergent micelles may be dissociated more easily than those formed by DDM as their non-ionic nature means that the only stabilizing forces are hydrophobic interactions which are weakened in the gas-phase, whilst DDM and b-OG micelles are also stabilized by hydrogenbonding [95].…”

Section: Detergent-based Reconstitution Methods For Native Msmentioning

confidence: 99%

“…Radford / Methods xxx (2018) xxx-xxxthe detergent micelle, i.e. harsher detergents require lower activation energies [97]. This means that different proteins solubilized in the same detergent may require different energies to be liberated into the gas phase.…”

Section: Detergent-based Reconstitution Methods For Native Msmentioning

confidence: 99%

“…This means that different proteins solubilized in the same detergent may require different energies to be liberated into the gas phase. It has been shown that the detergent used can influence significantly the observed charge states of a MP, and it has been posited that detergents may act as charge-carriers that result in nano-ESI droplets and MP ions with reduced charge (so the MP ions observed are more native-like) [97]. It should be noted, however, that it still remains the case that screening must be performed to determine empirically the optimal detergent to study a given MP by native MS [84].…”

Section: Detergent-based Reconstitution Methods For Native Msmentioning

confidence: 99%

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Mass spectrometry-enabled structural biology of membrane proteins

Calabrese

Radford

2018

Methods

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a b s t r a c tThe last $25 years has seen mass spectrometry (MS) emerge as an integral method in the structural biology toolkit. In particular, MS has enabled the structural characterization of proteins and protein assemblies that have been intractable by other methods, especially those that are large, heterogeneous or transient, providing experimental evidence for their structural organization in support of, and in advance of, high resolution methods. The most recent frontier conquered in the field of MS-based structural biology has been the application of established methods for studying water soluble proteins to the more challenging targets of integral membrane proteins. The power of MS in obtaining structural information has been enabled by advances in instrumentation and the development of hyphenated mass spectrometry-based methods, such as ion mobility spectrometry-MS, chemical crosslinking-MS and other chemical labelling/footprinting-MS methods. In this review we detail the insights garnered into the structural biology of membrane proteins by applying such techniques. Application and refinement of these methods has yielded unprecedented insights in many areas, including membrane protein conformation, dynamics, lipid/ligand binding, and conformational perturbations due to ligand binding, which can be challenging to study using other methods.

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“…Membrane proteins are liberated of detergent by collision-induced dissociation in an inert gas, after which their mass can be analyzed accurately (Barrera et al, 2008;Borysik and Robinson, 2012;Reading et al, 2015). Collision-induced dissociation does not only remove detergents but can also dissociate complexes and thus enable their compositional analysis (Laganowsky et al, 2014).…”

Section: Analysis Of the Molecular Mass Of Protein Complexesmentioning

confidence: 99%

Stoichiometry determination of macromolecular membrane protein complexes

Zilkenat

Grin

Wagner

2016

Biological Chemistry

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Gaining knowledge of the structural makeup of protein complexes is critical to advance our understanding of their formation and functions. This task is particularly challenging for transmembrane protein complexes, and grows ever more imposing with increasing size of these large macromolecular structures. The last 10 years have seen a steep increase in solved high-resolution membrane protein structures due to both new and improved methods in the field, but still most structures of large transmembrane complexes remain elusive. An important first step towards the structure elucidation of these difficult complexes is the determination of their stoichiometry, which we discuss in this review. Knowing the stoichiometry of complex components not only answers unresolved structural questions and is relevant for understanding the molecular mechanisms of macromolecular machines but also supports further attempts to obtain high-resolution structures by providing constraints for structure calculations.

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The Role of the Detergent Micelle in Preserving the Structure of Membrane Proteins in the Gas Phase

Cited by 126 publications

References 20 publications

Dimer interface of bovine cytochrome c oxidase is influenced by local posttranslational modifications and lipid binding

Dimer interface of bovine cytochrome c oxidase is influenced by local posttranslational modifications and lipid binding

Mass spectrometry-enabled structural biology of membrane proteins

Stoichiometry determination of macromolecular membrane protein complexes

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