METAL-MEDIATED HYDROLYSIS OF BIOLOGICAL PHOSPHATE-ESTERS - A CRITICALANALYSIS OF THE ESSENTIAL METAL-ION STOICHIOMETRY FOR MAGNESIUM-DEPENDENT NUCLEASE ACTIVATION

Metal-mediated hydrolysis of phosphate esters is a common catalytic pa thway in nucleic acid biochemistry. Two distinct models are principall y invoked in mechanistic discussions of these reactions for magnesium- dependent nuclease activation, namely, the one-versus two-metal-ion pa thways. By careful consideration of the available experimental data on a variety of enzymes (including the Klenow fragment of DNA pol I, exo nuclease III, ribonuclease H domains of HIV reverse transcriptase and the homologous Escherichia coli enzyme, avian sarcoma virus integrase, Eco RI and Eco RV restriction endonucleases, and ribozymes), it is de monstrated that the two-metal-ion paradigm cannot be substantiated as a bona fide mechanism on the basis of the published structural data. T he origins of the problems with this model are traced to fundamental i ssues in solution chemistry. These factors help to explain and resolve the often dichotomous results obtained from X-ray crystallography ver sus solution studies.