Kinetics of lipid mixing between bicelles and nanolipoprotein particles

Lai, Ginny; Forti, Kevin Muñoz; Renthal, Robert

doi:10.1016/j.bpc.2015.01.006

Cited by 11 publications

(20 citation statements)

References 42 publications

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“…We sought to demonstrate that the overall effect on the IMP is due to the gradual destruction of the ND as previous reports have shown the co-existence of NDs and diC6PC bicelles (Lai et al, 2015). Similarity between the NMR spectra of IMPs in titrated ND samples and in samples where MSP has been removed, confirms that the titration creates a smaller and more micellar environment for the IMP.…”

Section: Discussionmentioning

confidence: 99%

From Nanodiscs to Isotropic Bicelles: A Procedure for Solution Nuclear Magnetic Resonance Studies of Detergent-Sensitive Integral Membrane Proteins

et al. 2016

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Summary Nanodiscs and isotropic bicelles are promising membrane mimetics in the field of solution NMR spectroscopy of integral membrane proteins (IMPs). Despite varied challenges to solution NMR studies of IMPs, we attribute the paucity of solution NMR structures in these environments to the inability of diverse IMPs to withstand detergent treatment during standard nanodisc and bicelle preparations. Here, we present a strategy that creates small isotropic bicelles from IMPs co-translationally embedded in large nanodiscs using cell-free expression. Our results demonstrate appreciable gains in NMR spectral quality while preserving lipid-IMP contacts. We validate the approach on the detergent sensitive LspA, which finally allowed us to perform high quality triple resonance NMR experiments for structural studies. Our strategy of producing bicelles from nanodiscs comprehensively avoids detergent during expression and preparation and is suitable for solution NMR spectroscopy of lipid-IMPs complexes.

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Section: Discussionmentioning

confidence: 99%

From Nanodiscs to Isotropic Bicelles: A Procedure for Solution Nuclear Magnetic Resonance Studies of Detergent-Sensitive Integral Membrane Proteins

et al. 2016

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“…448 Lipid transfer was monitored by gradual disappearance of FRET, and Arrhenius dependence of rate revealed high activation energy, 28 kcal/mol, measured between 20.5 °C and 33.2 °C. FRET dependence on DHPC concentration was also studied in order to take into account this side reaction.…”

Section: Application Of Nanodiscs In Functional Studies Of Membranmentioning

confidence: 99%

“…449 This feature, together with the similar high activation barriers observed in fusion pores formation, suggests the same mechanism for lipid exchange between Nanodiscs and bicelles. 448 …”

Section: Application Of Nanodiscs In Functional Studies Of Membranmentioning

confidence: 99%

Nanodiscs in Membrane Biochemistry and Biophysics

2017

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Membrane proteins play a most important part in metabolism, signaling, cell motility, transport, development, and many other biochemical and biophysical processes which constitute fundamentals of life on molecular level. Detailed understanding of these processes is necessary for the progress of life sciences and biomedical applications. Nanodiscs provide a new and powerful tool for a broad spectrum of biochemical and biophysical studies of membrane proteins and are commonly acknowledged as an optimal membrane mimetic system which provides control over size, composition and specific functional modifications on nanometer scale. In this review we attempted to combine a comprehensive list of various applications of Nanodisc technology with systematic analysis of the most attractive features of this system and advantages provided by Nanodiscs for structural and mechanistic studies of membrane proteins.

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“…Co-translation of the NLP platform has also proven useful for functional characterization using multiple biochemical and biophysical techniques (Blanchette et al, 2009b; Gao et al, 2011; He et al, 2013; Lai et al, 2015; Cleveland et al, 2018). Atomic force microscopy (AFM), EM, and FCS have been critical in general to facilitate characterization of the nanoparticles and the functional attributes of the embedded membrane proteins.…”

Section: Summary and Future Perspectivesmentioning

confidence: 99%

Cell-Free Co-Translational Approaches for Producing Mammalian Receptors: Expanding the Cell-Free Expression Toolbox Using Nanolipoproteins

Shelby

Dang

et al. 2019

Front. Pharmacol.

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Membranes proteins make up more than 60% of current drug targets and account for approximately 30% or more of the cellular proteome. Access to this important class of proteins has been difficult due to their inherent insolubility and tendency to aggregate in aqueous solutions. Understanding membrane protein structure and function demands novel means of membrane protein production that preserve both their native conformational state as well as function. Over the last decade, cell-free expression systems have emerged as an important complement to cell-based expression of membrane proteins due to their simple and customizable experimental parameters. One approach to overcome the solubility and stability limitations of purified membrane proteins is to support them in stable, native-like states within nanolipoprotein particles (NLPs), aka nanodiscs. This has become common practice to facilitate biochemical and biophysical characterization of proteins of interest. NLP technology can be easily coupled with cell-free systems to achieve functional membrane protein production for this purpose. Our approach involves utilizing cell-free expression systems in the presence of NLPs or using co-translation techniques to perform one-pot expression and self-assembly of membrane protein/NLP complexes. We describe how cell-free reactions can be modified to render control over nanoparticle size and monodispersity in support of membrane protein production. These modifications have been exploited to facilitate co-expression of full-length functional membrane proteins such as G-protein-coupled receptors (GPCRs) and receptor tyrosine kinases (RTKs). In particular, we summarize the state of the art in NLP-assisted cell-free coexpression of these important classes of membrane proteins as well as evaluate the advances in and prospects for this technology that will drive drug discovery against these targets. We conclude with a prospective on the use of NLPs to produce as well as deliver functional mammalian membrane-bound proteins for a range of applications.

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Kinetics of lipid mixing between bicelles and nanolipoprotein particles

Cited by 11 publications

References 42 publications

From Nanodiscs to Isotropic Bicelles: A Procedure for Solution Nuclear Magnetic Resonance Studies of Detergent-Sensitive Integral Membrane Proteins

From Nanodiscs to Isotropic Bicelles: A Procedure for Solution Nuclear Magnetic Resonance Studies of Detergent-Sensitive Integral Membrane Proteins

Nanodiscs in Membrane Biochemistry and Biophysics

Cell-Free Co-Translational Approaches for Producing Mammalian Receptors: Expanding the Cell-Free Expression Toolbox Using Nanolipoproteins

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