title: Identification of (eta6-arene)ruthenium(II) protein binding sites in E. coli cells by combined multidimensional liquid chromatography and ESI tandem mass spectrometry: specific binding of [(eta6-p-cymene)RuCl2 (DMSO)] to stress-regulated proteins and to helicases.
abstract: An automated multidimensional protein identification technology, which combines biphasic liquid chromatography with electrospray ionisation tandem mass spectrometry (MS/MS), was employed to analyse tryptic peptides from Escherichia coli cells treated with the antiproliferation agent [(eta(6)-p-cymene)RuCl(2)(DMSO)], where DMSO is dimethyl sulfoxide. MS/MS spectra were recorded for molecular ions generated by neutral loss of p-cymene from intensive peptide ions coordinated by the (eta(6)-p-cymene)Ru(II) fragment. Matching of the MS/MS spectra of the ruthenated peptides to spectra of proteins in the E. coli database enabled the identification of five protein targets for [(eta(6)-p-cymene)RuCl(2)(DMSO)]. One of these is the constitutive cold-shock protein cspC, which regulates the expression of genes encoding stress-response proteins, and three of the other targets, ppiD, osmY and sucC, are proteins of the latter type. The DNA damage-inducible helicase dinG was likewise established as a protein target. Aspartate carboxylate functions were identified as the probable Ru binding sites in cspC, ppiD and dinG, and threonine and lysine side chains in osmY and sucC, respectively.
authors.affiliation: Lehrstuhl für Analytische Chemie, Ruhr-Universität Bochum, 44780 Bochum, Germany.
authors.forename: Joanna
authors.initials: J
authors.lastname: Will
authors.forename: Andreas
authors.initials: A
authors.lastname: Kyas
authors.forename: William S
authors.initials: WS
authors.lastname: Sheldrick
authors.forename: Dirk
authors.initials: D
authors.lastname: Wolters
mesh_terms: D000970:Antineoplastic Agents / Q000737:chemistry; D001665:Binding Sites; D002853:Chromatography, Liquid; D004265:DNA Helicases; D004121:Dimethyl Sulfoxide; D029968:Escherichia coli Proteins / Q000378:metabolism*; D006360:Heat-Shock Proteins; D040901:Proteomics / Q000379:methods*; D017975:Ruthenium Compounds / Q000737:chemistry*; D021241:Spectrometry, Mass, Electrospray Ionization; D053719:Tandem Mass Spectrometry
chemical_list: D000970:Antineoplastic Agents; D029968:Escherichia coli Proteins; D006360:Heat-Shock Proteins; D017975:Ruthenium Compounds; D004265:DNA Helicases; D004121:Dimethyl Sulfoxide