Native MS Analysis of Bacteriorhodopsin and an Empty Nanodisc by Orthogonal Acceleration Time-of-Flight, Orbitrap and Ion Cyclotron Resonance
| Autor: | Juraj Svitel, Han Xu, Jennifer L. Lippens, Joseph A. Loo, Robert J. M. Kurzeja, Iain Campuzano, Dhanashri Bagal, Paul D. Schnier, Huilin Li |
|---|---|
| Rok vydání: | 2016 |
| Předmět: |
Halobacterium salinarum
Models Molecular 0301 basic medicine Spectrum analyzer Protein Conformation Cyclotron Analytical chemistry Mass spectrometry Orbitrap 01 natural sciences Article Mass Spectrometry Analytical Chemistry law.invention 03 medical and health sciences Acceleration Glucosides Purple Membrane law Micelles Nanodisc Fourier Analysis biology Chemistry 010401 analytical chemistry Bacteriorhodopsin Cyclotrons Nanostructures 0104 chemical sciences 030104 developmental biology Bacteriorhodopsins biology.protein Ion cyclotron resonance |
| Zdroj: | Analytical Chemistry. 88:12427-12436 |
| ISSN: | 1520-6882 0003-2700 |
| Popis: | Over the past two decades orthogonal acceleration time-of-flight has been the de facto analyzer of choice for solution and membrane soluble protein native mass spectrometry (MS) studies; this however is gradually changing. Here we compare three MS instruments, the Q-ToF, the Orbitrap and the FT-ICR to analyze, under native instrument and buffer conditions, the 7-transmembrane helical protein bacteriorhodopsin-octylglucoside micelle complex and the empty nanodisc (MSP1D1-Nd) using both MS and tandem-MS modes of operation. Bacteriorhodopsin can be released from the octylglucoside-micelle efficiently on all three instruments (MS-mode of operation) producing a narrow charge state distribution (z = 8+ to 10+) by either increasing the source lens or collision cell (or HCD) voltages. A lower center-of-mass collision energy (0.20–0.41 eV) is required for optimal bacteriorhodopsin liberation on the FT-ICR, in comparison to the Q-ToF and Orbitrap instruments (0.29–2.47 eV). The empty MSP1D1-Nd can be measured with relative ease on a three instruments, resulting in a highly complex spectrum of overlapping, polydisperse charge state; a consequence of varying levels of phospholipid incorporation. There is a measurable difference in MSP1D1-Nd charge state distribution (z = 15+ to 26+), average molecular weight (141.7 to 169.6 kDa) and phospholipid incorporation number (143 to 184) under low activation conditions. Utilizing tandem-MS, bacteriorhodopsin can be effectively liberated from the octylglucoside-micelle by collisional (Q-ToF and FT-ICR) or continuous IRMPD activation (FT-ICR). MSP1D1-Nd spectral complexity can also be significantly reduced by tandem-MS (Q-ToF and FT-ICR) followed by mild collisional or continuous IRMPD activation, resulting in a spectrum in which the charge state and phospholipid incorporation levels can easily be determined. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|