title: Proteomic analysis of complexes formed by human topoisomerase I.
abstract: Human topoisomerase I is a nuclear enzyme that catalyses DNA relaxation and phosphorylation of SR proteins. Topoisomerase I participates in several protein-protein interactions. We performed a proteomic analysis of protein partners of topoisomerase I. Two methods were applied to proteins of the nuclear extract of HeLa cells: a co-immunoprecipitation and an affinity chromatography combined with mass spectrometry. Complexes formed by topoisomerase I with its protein partners were immunoprecipitated by scleroderma anti-topoisomerase I antibodies. To identify binding sites for the protein partners, baits corresponding to fragments of topoisomerase I were constructed and used in the affinity chromatography. The N-terminal domain and the cap region of the core domain appeared to be the main regions that bound proteins. We identified 36 nuclear proteins that were associated with topoisomerase I. The proteins were mainly involved in RNA metabolism. We found 29 new and confirmed 7 previously identified protein partners of topoisomerase I. More than 40% proteins that associate with the cap region contain two closely spaced RRM domains. Docking calculations identified the RRM domains as a possible site for the interaction of these proteins with the cap region.
authors.affiliation: Institute of Biochemistry, Warsaw University, ul. Miecznikowa 1, 02-096 Warszawa, Poland.
authors.forename: Alicja
authors.initials: A
authors.lastname: Czubaty
authors.forename: Agnieszka
authors.initials: A
authors.lastname: Girstun
authors.forename: Barbara
authors.initials: B
authors.lastname: Kowalska-Loth
authors.forename: Agata M
authors.initials: AM
authors.lastname: Trzcińska
authors.forename: Elzbieta
authors.initials: E
authors.lastname: Purta
authors.forename: Alicja
authors.initials: A
authors.lastname: Winczura
authors.forename: Wojciech
authors.initials: W
authors.lastname: Grajkowski
authors.forename: Krzysztof
authors.initials: K
authors.lastname: Staroń
mesh_terms: D004264:DNA Topoisomerases, Type I / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D047468:Immunoprecipitation; D009687:Nuclear Proteins / Q000378:metabolism*; D011487:Protein Conformation; D040901:Proteomics; D012313:RNA / Q000378:metabolism; D012261:Ribonucleoproteins / Q000032:analysis / Q000378:metabolism
chemical_list: D009687:Nuclear Proteins; D012261:Ribonucleoproteins; D012313:RNA; D004264:DNA Topoisomerases, Type I