Michael Marty - Ultra-thin layer MALDI mass spectrometry of membrane proteins in nanodiscs.

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  Publication Details (including relevant citation   information): Marty,Michael T., Das,Aditi,   Sligar,Stephen G., Anal. Bioanal. Chem.,   2012, 402 (2), pp 721-729

  Abstract: Nanodiscs have become a leading   technol. to solubilize membrane proteins for biophys., enzymic,   and structural investigations. Nanodiscs are nanoscale, discoidal   lipid bilayers surrounded by an amphipathic membrane scaffold   protein (MSP) belt. A variety of anal. tools has been applied to   membrane proteins in nanodiscs, including several recent mass   spectrometry studies. Mass spectrometry of full-length proteins   is an important technique for analyzing protein modifications,   for structural studies, and for identification of proteins   present in binding assays. However, traditional matrix-assisted   laser desorption/ionization-time-of-flight (MALDI-TOF) mass   spectrometry methods for analyzing full-length membrane proteins   solubilized in nanodiscs are limited by strong signal from the   MSP belt and weak signal from the membrane protein inside the   nanodisc. Herein, we show that an optimized ultra-thin layer   MALDI sample prepn. technique dramatically enhances the membrane   protein signal and nearly completely eliminates the MSP signal.   First-shot MALDI and MALDI imaging are used to characterize the   spots formed by the ultra-thin layer method. Furthermore, the   membrane protein enhancement and MSP suppression are shown to be   independent of the type of membrane protein and are applicable to   mixts. of membrane proteins in nanodiscs. FigureA model of   cytochrome P 450 3A4 (CYP 3A4) in a sol. lipid bilayer nanodisc   is shown (left). Membrane proteins in nanodiscs demonstrate poor   spectra with MALDI mass spectrometry using the conventional   dried-drop method (middle). The ultra-thin layer method shows   remarkable enhancement of membrane protein signal and nearly   complete suppression of the membrane scaffold protein belt (MSP)   signal (right). [on SciFinder(R)]

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