title: Dual-targeted labeling of proteins using cysteine and selenomethionine residues.
abstract: A new strategy for dual site-selective labeling of proteins that uses metabolically incorporated selenomethionine as a target for covalent modification by iodoacetamide derivatives, forming selenonium salts, is described. In the absence of free cysteine, labeling is specific and efficient. Dual-targeted labeling of a protein can be achieved with combinations of unique cysteine and methionine residues, if the cysteine is labeled first with a maleimide or another reagent that does not react with the selenomethionine. The method should be useful in biophysical applications such as fluorescence energy transfer.
authors.affiliation: Department of Chemistry, University of Waterloo, Waterloo, Ont., Canada N2L 3G1.
authors.forename: Shenhui
authors.initials: S
authors.lastname: Lang
authors.forename: Donald E
authors.initials: DE
authors.lastname: Spratt
authors.forename: J Guy
authors.initials: JG
authors.lastname: Guillemette
authors.forename: Michael
authors.initials: M
authors.lastname: Palmer
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry; D002147:Calmodulin / Q000737:chemistry / Q000235:genetics; D003545:Cysteine / Q000737:chemistry*; D004735:Energy Transfer; D004926:Escherichia coli / Q000378:metabolism; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000737:chemistry*; D006460:Hemolysin Proteins; D007460:Iodoacetamide / Q000737:chemistry; D008301:Maleimides / Q000737:chemistry; D016297:Mutagenesis, Site-Directed; D009282:Naphthalenesulfonates / Q000737:chemistry; D011506:Proteins / Q000737:chemistry*; D012645:Selenomethionine / Q000737:chemistry*
chemical_list: C032162:AEDANS-calmodulin; D001426:Bacterial Proteins; C039092:CAMP protein, Streptococcus; D002147:Calmodulin; D005456:Fluorescent Dyes; D006460:Hemolysin Proteins; D008301:Maleimides; D009282:Naphthalenesulfonates; D011506:Proteins; C043592:maleimide; D012645:Selenomethionine; D003545:Cysteine; D007460:Iodoacetamide