TY - JOUR
T1 - Amphipathic polymers enable the study of functional membrane proteins in the gas phase
AU - Leney, Aneika C.
AU - McMorran, Lindsay M.
AU - Radford, Sheena E.
AU - Ashcroft, Alison E.
PY - 2012/11/20
Y1 - 2012/11/20
N2 - Membrane proteins are notoriously challenging to analyze using mass spectrometry (MS) because of their insolubility in aqueous solution. Current MS methods for studying intact membrane proteins involve solubilization in detergent. However, detergents can destabilize proteins, leading to protein unfolding and aggregation, or resulting in inactive entities. Amphipathic polymers, termed amphipols, can be used as a substitute for detergents and have been shown to enhance the stability of membrane proteins. Here, we show the utility of amphipols for investigating the structural and functional properties of membrane proteins using electrospray ionization mass spectrometry (ESI-MS). The functional properties of two bacterial outer-membrane β-barrel proteins, OmpT and PagP, in complex with the amphipol A8-35 are demonstrated, and their structural integrities are confirmed in the gas phase using ESI-MS coupled with ion mobility spectrometry (IMS). The data illustrate the power of ESI-IMS-MS in separating distinct populations of amphipathic polymers from the amphipol-membrane complex while maintaining a conformationally "nativelike" membrane protein structure in the gas phase. Together, the data indicate the potential importance and utility of amphipols for the analysis of membrane proteins using MS.
AB - Membrane proteins are notoriously challenging to analyze using mass spectrometry (MS) because of their insolubility in aqueous solution. Current MS methods for studying intact membrane proteins involve solubilization in detergent. However, detergents can destabilize proteins, leading to protein unfolding and aggregation, or resulting in inactive entities. Amphipathic polymers, termed amphipols, can be used as a substitute for detergents and have been shown to enhance the stability of membrane proteins. Here, we show the utility of amphipols for investigating the structural and functional properties of membrane proteins using electrospray ionization mass spectrometry (ESI-MS). The functional properties of two bacterial outer-membrane β-barrel proteins, OmpT and PagP, in complex with the amphipol A8-35 are demonstrated, and their structural integrities are confirmed in the gas phase using ESI-MS coupled with ion mobility spectrometry (IMS). The data illustrate the power of ESI-IMS-MS in separating distinct populations of amphipathic polymers from the amphipol-membrane complex while maintaining a conformationally "nativelike" membrane protein structure in the gas phase. Together, the data indicate the potential importance and utility of amphipols for the analysis of membrane proteins using MS.
UR - http://www.scopus.com/inward/record.url?scp=84869398484&partnerID=8YFLogxK
U2 - 10.1021/ac302223s
DO - 10.1021/ac302223s
M3 - Article
C2 - 23072351
AN - SCOPUS:84869398484
SN - 0003-2700
VL - 84
SP - 9841
EP - 9847
JO - Analytical Chemistry
JF - Analytical Chemistry
IS - 22
ER -