title: In vitro and in vivo chemical labeling of ribosomal proteins: a quantitative comparison.
abstract: Thioimidates have emerged as reagents for probing the protein structure, folding, and interactions under physiological conditions. The same properties that give thioimidates biological relevance make these molecules ideal candidates for use in vivo. Through labeling of ribosomal proteins, we have quantified the in vivo and in vitro reactivity of two thioimidates: S-methylthioacetimidate (SMTA) and a novel, charge-carrying analogue, S-sulfethylthioacetimidate (SSETA). In vitro experiments demonstrate that both amidinating reagents can probe the protein structure. Under comparable in vivo conditions, SMTA is found to be membrane-permeable while SSETA is not. The use of mass spectrometry with permeant and impermeant thioimidates promises insights into the membrane topology and protein structure in the native environment.
authors.affiliation: Department of Chemistry, Indiana University, Bloomington, Indiana 47405-7000, United States.
authors.forename: Ethan G
authors.initials: EG
authors.lastname: Jaffee
authors.forename: Matthew A
authors.initials: MA
authors.lastname: Lauber
authors.forename: William E
authors.initials: WE
authors.lastname: Running
authors.forename: James P
authors.initials: JP
authors.lastname: Reilly
mesh_terms: D002463:Cell Membrane Permeability; D048168:Escherichia coli K12 / Q000166:cytology; D007096:Imidoesters / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism; D012269:Ribosomal Proteins / Q000737:chemistry*; D013194:Staining and Labeling / Q000379:methods*
chemical_list: D007096:Imidoesters; D012269:Ribosomal Proteins; C576674:S-methylthioacetimidate