Characterization of Mg2+ Binding to the DNA Repair Protein Apurinic/Apyrimidic Endonuclease 1 (APE1) via Solid-State 25Mg NMR Spectroscopy

摘要

Apurinic/apyrimidinic endonuclease 1 (APE1), a member of the divalent cation-dependent phosphoesterase superfamily of proteins that retain the conserved four-layered α/β-sandwich structural core, is an essential protein that functions as part of base excision repair to remove mutagenic and cytotoxic abasic sites from DNA. Using low temperature solid-state ²⁵Mg NMR spectroscopy and various mutants of APE1, we demonstrate that Mg²⁺ binds to APE1 and a functional APE1-substrate DNA complex with an overall stoichiometry with respect to Mg²⁺ of one per mole of APE1 as predicted by the X-ray work of Tainer and coworkers. However, the NMR spectra show that the single Mg²⁺ site is disordered. We discuss the probable reasons for the disorder at the Mg²⁺ binding site. The most likely source of this disorder is arrangement of the protein ligands about the Mg²⁺ (cis and trans isomers). The existence of these isomers reinforces the notion of the plasticity of the metal binding site within APE1.