Proteomics on Full-Length Membrane Proteins Using Mass Spectrometry†
journal contributionposted on 21.03.2000, 00:00 by Johannes le Coutre, Julian P. Whitelegge, Adrian Gross, Eric Turk, Ernest M. Wright, H. Ronald Kaback, Kym F. Faull
A general technique has been developed that allows rapid mass spectrometric analysis of full-length membrane proteins [Whitelegge, J. P., le Coutre, J., et al. (1999) Proc. Natl. Acad. Sci. U.S.A. 96, 10695−10698]. Using in-line HPLC electrospray ionization mass spectrometry (LC−MS), different native and recombinant bacterial membrane proteins of up to 61 kDa are characterized. Mass spectrometric data of four entirely different membrane proteins from three bacterial organisms, two transporters, a channel, and a porin protein are presented. In addition to determination of the molecular mass with an accuracy of ±0.01%, the technique monitors alkylation or oxidation of single Cys residues and errors in deduced amino acid sequences. Finally, using in-line LC−MS, unknown proteins can be identified from solubilized Escherichia coli membranes without prior purification.