%0 Generic
%A Jagoda, Malgorzata
%D 2005
%F heidok:5930
%K Phosphorylierungskatalysatoren , DNA-Polymerase , Phosphoryltransfer , Makrocyklen , Dikupfer(II)-Komplexphosphoryltransfer , low-molecular-weight , DNA  polymerase , dicopper(II) complex
%R 10.11588/heidok.00005930
%T Bioinspired Catalysts for Phosphoryl Transfer Reactions
%U https://archiv.ub.uni-heidelberg.de/volltextserver/5930/
%X Phosphoryl transfer reactions are ubiquitous in biology and are usually catalyzed by metalloenzymes. For a number of phosphoryltransfer enzymes, including the exonuclease subunit of DNA  polymerase I, a mechanism involving two metal ions and double Lewis-acid activation of the substrate, combined with leaving group stabilisation, has been proposed. Aim of this thesis was the study of low-molecular-weight models of phosphoryl transfer metalloenzymes. The focus was in particular on the macrocyclic dicopper(II) complex LCu2, which is the first transesterification catalyst for highly inert dialkyl phosphates and a mimic of the exonuclease subunit of DNA polymerase I. For the first time, a crystal structure of LCu2 with a coordinated phosphodiester was obtained. [LCu2(1,3-µ-DMP)(NO3)](NO3)2 (2) contains a 1,3- bridging phosphodiester.  By extensive UV-Vis and pH metric titrations, the metal complex species present in methanolic LCu2/DMP solution have been identified. In particular it has been shown, that [LCu2(DMP)(OCH3)]2+ is the active species and that coordinated methanolate (and not free methanol or methanolate) is the nucleophile which attacks the P-atom of the substrate. DFT calculations (in cooperation with the group of Prof. Smith, IWR Heidelberg) confirm that the 1,3-DMP bridged complex is the dominant species in solution.  By extensive kinetic studies including rate dependence on catalyst concentration, saturation kinetics, pH dependence on rate and dependence on rate on substrate structure, solid experimental data have been obtained which support the proposal of a reaction mechanism related to exonuclease subunit of DNA polymerase I.  The applicability of LCu2 as a catalyst for the phosphorylation of various alcohols by dimethyl phosphate has been evaluated. A unique functional group tolerance of the catalyst and selectivity for dialkyl phosphates over alkyl carboxylates is observed. Screening of ATP hydrolysis by Zr(IV) and Eu(III) complexes was examined using a robotic liquid-handling workstation. It was found that a phosphate detection assay using molybdate is compatible with the presence of these high-valent metal ions. ATPase activity of the metal is modulated by the ligands, and the most efficient catalyst is the Zr complex of the tetracarboxylate ligand L6 at both pH 4 and pH 7. This is in contrast to Zr promoted DNA hydrolysis where polycarboxylate ligands have been shown to quench strongly the phosphoesterase activity of the metal ion.