Abstract
Native liquid extraction surface analysis (LESA) mass spectrometry allows direct analysis of folded proteins and protein complexes from biological substrates, such as dried blood spots and thin tissue sections, by use of native-like extraction/ionization solvents. Previously, we have demonstrated native LESA mass spectrometry of folded proteins up to 16 kDa as well as the 64 kDa hemoglobin tetramer, from mouse tissues. With denaturing LESA solvents, the highest mass protein detected in tissue to date is ∼37 kDa. Here, we demonstrate native LESA mass spectrometry by use of a Q Exactive UHMR Hybrid Quadrupole-Orbitrap (QE-UHMR) mass spectrometer, pushing the upper mass limit of proteins detected in tissue to >70 kDa. Moreover, a protein trimer of 42 kDa was detected and its stoichiometry confirmed by higher energy collision dissociation (HCD). The benefits of inclusion of detergents in the LESA sampling solvent are also demonstrated.
Highlights
Native mass spectrometry is emerging as a powerful tool for the analysis of protein complexes and assemblies, providing structural and stoichiometric insights.[1]
We have shown that Liquid extraction surface analysis (LESA) MS is suited to the in situ analysis of intact proteins from substrates including thin tissue sections,[8] dried blood spots,[9] and bacterial colonies growing on agar.[10]
We present native LESA MS of thin tissue sections of rat brain and rat kidney on an QE-UHMR mass spectrometer, pushing the upper mass limit of proteins detected from tissue to >70 kDa
Summary
Native mass spectrometry is emerging as a powerful tool for the analysis of protein complexes and assemblies, providing structural and stoichiometric insights.[1]. We have shown that LESA MS is suited to the in situ analysis of intact proteins from substrates including thin tissue sections,[8] dried blood spots,[9] and bacterial colonies growing on agar.[10] The majority of work to date has made use of denaturing solvents, and typically proteins up to mass 20 kDa are detected, proteins up to 37 kDa have been observed when high-field asymmetric waveform ion mobility (FAIMS) has been employed[11]. We have demonstrated native LESA mass spectrometry, in which folded proteins and protein assemblies are extracted directly from their substrate, on a Q-TOF mass spectrometer.[12,13] Initial work focused on protein standards dried onto glass substrates. We show the benefits of inclusion of tetraethylene glycol monooctyl ether (C8E4) detergent in the LESA sampling/electrospray solvent
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