Conformational analysis of membrane proteins in phospholipid bilayer nanodiscs by hydrogen exchange mass spectrometry

TY - JOUR

T1 - Conformational analysis of membrane proteins in phospholipid bilayer nanodiscs by hydrogen exchange mass spectrometry

AU - Hebling, Christine M

AU - Morgan, Christopher R

AU - Stafford, Darrel W

AU - Jorgenson, James W

AU - Rand, Kasper Dyrberg

AU - Engen, John R

PY - 2010/7/1

Y1 - 2010/7/1

N2 - The study of membrane protein structure and enzymology has traditionally been hampered by the inherent insolubility of membrane proteins in aqueous environments and experimental challenges in emulating an in vivo lipid environment. Phospholipid bilayer nanodiscs have recently been shown to be of great use for the study of membrane proteins since they offer a controllable, stable, and monodisperse model membrane with a nativelike lipid bilayer. Here we report the integration of nanodiscs with hydrogen exchange (HX) mass spectrometry (MS) experiments, thereby allowing for analysis of the native conformation of membrane proteins. γ-Glutamyl carboxylase (GGCX), an ∼94 kDa transmembrane protein, was inserted into nanodiscs and labeled with deuterium oxide under native conditions. Analytical parameters including sample-handling and chromatographic separation were optimized to measure the incorporation of deuterium into GGCX. Coupling nanodisc technology with HX MS offers an effective approach for investigating the conformation and dynamics of membrane proteins in their native environment and is therefore capable of providing much needed insight into the function of membrane proteins.

AB - The study of membrane protein structure and enzymology has traditionally been hampered by the inherent insolubility of membrane proteins in aqueous environments and experimental challenges in emulating an in vivo lipid environment. Phospholipid bilayer nanodiscs have recently been shown to be of great use for the study of membrane proteins since they offer a controllable, stable, and monodisperse model membrane with a nativelike lipid bilayer. Here we report the integration of nanodiscs with hydrogen exchange (HX) mass spectrometry (MS) experiments, thereby allowing for analysis of the native conformation of membrane proteins. γ-Glutamyl carboxylase (GGCX), an ∼94 kDa transmembrane protein, was inserted into nanodiscs and labeled with deuterium oxide under native conditions. Analytical parameters including sample-handling and chromatographic separation were optimized to measure the incorporation of deuterium into GGCX. Coupling nanodisc technology with HX MS offers an effective approach for investigating the conformation and dynamics of membrane proteins in their native environment and is therefore capable of providing much needed insight into the function of membrane proteins.

U2 - 10.1021/ac100962c

DO - 10.1021/ac100962c

M3 - Journal article

C2 - 20518534

SN - 0003-2700

VL - 82

SP - 5415

EP - 5419

JO - Industrial And Engineering Chemistry Analytical Edition

JF - Industrial And Engineering Chemistry Analytical Edition

IS - 13

ER -