In vivo incorporation of selenomethionine in proteins using Pseudomonas aeruginosa as expression host:: case study -: the outer membrane receptor FpvA

The number of protein structures solved using multiwavelength anomalous diffraction methods coupled with selenomethionine substitution has grown dramatically over the last years. We show using the outer membrane pyoverdin receptor FpvA that Pseudomonas aeruginosa can be used for producing proteins with a high level of selenomethionine incorporation. To circumvent problems encountered with mass spectroscopy analysis of purified membrane proteins, in-gel trypsin digestion of FpvA coupled with MALDI mass spectrometry analysis of the resulting peptides was used to determine the extent of selenomethionine incorporation. Selenomethionine incorporation greater than 95% was achieved using P. aeruginosa as an overexpression system. (C) 2004 Elsevier Inc. All rights reserved.