title: Clofarabine targets the large subunit (α) of human ribonucleotide reductase in live cells by assembly into persistent hexamers.
abstract: Clofarabine (ClF) is a drug used in the treatment of leukemia. One of its primary targets is human ribonucleotide reductase (hRNR), a dual-subunit, (α(2))(m)(β(2))(n), regulatory enzyme indispensable in de novo dNTP synthesis. We report that, in live mammalian cells, ClF targets hRNR by converting its α-subunit into kinetically stable hexamers. We established mammalian expression platforms that enabled isolation of functional α and characterization of its altered oligomeric associations in response to ClF treatment. Size exclusion chromatography and electron microscopy documented persistence of in-cell-assembled-α(6). Our data validate hRNR as an important target of ClF, provide evidence that in vivo α's quaternary structure can be perturbed by a nonnatural ligand, and suggest small-molecule-promoted, persistent hexamerization as a strategy to modulate hRNR activity. These studies lay foundations for documentation of RNR oligomeric state within a cell.
authors: Department of Chemistry, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors: Yimon
authors: Y
authors: Aye
authors: Edward J
authors: EJ
authors: Brignole
authors: Marcus J C
authors: MJ
authors: Long
authors: Johnathan
authors: J
authors: Chittuluru
authors: Catherine L
authors: CL
authors: Drennan
authors: Francisco J
authors: FJ
authors: Asturias
authors: JoAnne
authors: J
authors: Stubbe
mesh_terms: D000227:Adenine Nucleotides / Q000494:pharmacology*; D001087:Arabinonucleosides / Q000494:pharmacology*; D002470:Cell Survival; D000077866:Clofarabine; D006801:Humans; D007700:Kinetics; D008099:Liver / Q000166:cytology* / Q000187:drug effects* / Q000201:enzymology; D015394:Molecular Structure; D011487:Protein Conformation / Q000187:drug effects; D055503:Protein Multimerization / Q000187:drug effects*; D021122:Protein Subunits / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D012264:Ribonucleotide Reductases / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000378:metabolism
chemical_list: D000227:Adenine Nucleotides; D001087:Arabinonucleosides; D021122:Protein Subunits; D000077866:Clofarabine; D012264:Ribonucleotide Reductases