Automated SDS Depletion for Mass Spectrometry of Intact Membrane Proteins though Transmembrane Electrophoresis

Membrane proteins are underrepresented
in proteome analysis platforms
because of their hydrophobic character, contributing to decreased
solubility. Sodium dodecyl sulfate is a favored denaturant in proteomic
workflows, facilitating cell lysis and protein dissolution; however,
SDS impedes MS detection and therefore must be removed prior to analysis.
Although strategies exist for SDS removal, they provide low recovery,
purity, or reproducibility. Here we present a simple automated device,
termed transmembrane electrophoresis (TME), incorporating the principles
of membrane filtration, but with an applied electric current to ensure
near-complete (99.9%) removal of the surfactant, including protein-bound
SDS. Intact proteins are recovered in solution phase in high yield
(90–100%) within 1 h of operation. The strategy is applied
to protein standards and proteome mixtures, including an enriched
membrane fraction from E. coli, resulting in quality
MS spectra free of SDS adducts. The TME platform is applicable to
both bottom-up MS/MS as well as LC–ESI–MS analysis of
intact proteins. SDS-depleted fractions reveal a similar number of
protein identifications (285) compared wit a non-SDS control (280),
being highly correlated in terms of protein spectral counts. This
fully automated approach to SDS removal presents a viable tool for
proteome sample processing ahead of MS analysis. Data are available
via ProteomeXchange, identifier PXD003941.