Identification of uranyl binding proteins from human kidney-2 cell extracts by immobilized uranyl affinity chromatography and mass spectrometry.
To improve our knowledge on protein targets of uranyl ion (UO(2)(2+)), we set up a proteomic strategy based on immobilized metal-affinity chromatography (IMAC). The successful enrichment of UO(2)(2+)-interacting proteins from human kidney-2 (HK-2) soluble cell extracts was obtained using an ion-exchange chromatography followed by a dedicated IMAC process previously described and designed for the uranyl ion. By mass spectrometry analysis we identified 64 proteins displaying varied functions. The use of a computational screening algorithm along with the particular ligand-based properties of the UO(2)(2+) ion allowed the analysis and categorization of the protein collection. This profitable approach demonstrated that most of these proteins fulfill criteria which could rationalize their binding to the UO(2)(2+)-loaded phase. The obtained results enable us to focus on some targets for more in-depth studies and open new insights on its toxicity mechanisms at molecular level.
Dedieu
Alain
A
CEA/IBEB/SBTN, F-30207 Bagnols sur Cèze, France.
Bérenguer
Frédéric
F
Basset
Christian
C
Prat
Odette
O
Quéméneur
Eric
E
Pible
Olivier
O
Vidaud
Claude
C
D000465:Algorithms; D002457:Cell Extracts / Q000737:chemistry*; D002460:Cell Line; D055598:Chemical Phenomena; D002846:Chromatography, Affinity / Q000379:methods*; D003198:Computer Simulation; D015180:Electrophoresis, Gel, Two-Dimensional; D006801:Humans; D007477:Ions / Q000378:metabolism; D007668:Kidney / Q000166:cytology*; D013058:Mass Spectrometry / Q000379:methods*; D011485:Protein Binding; D011506:Proteins / Q000032:analysis / Q000378:metabolism*; D020543:Proteome / Q000032:analysis / Q000378:metabolism; D040901:Proteomics / Q000379:methods; D014501:Uranium / Q000378:metabolism*
D002457:Cell Extracts; D007477:Ions; D011506:Proteins; D020543:Proteome; D014501:Uranium