Expression, Purification, and Characterization of Recombinant Saccharomyces Cerevisiae Adenosine Kinase

Adenosine kinase (AK), a key enzyme in the regulation of the cellular concentrations of adenosine (A), is an important physiological effector of many cells and tissues. In this article, we reported that ak, which encoded adenosine kinase, was cloned from Saccharomyces cerevisiae, sequenced, and overexpressed in E. coli using the pET16b expression system, and the recombinant protein was purified to apparent homogeneity using conventional protein purification techniques. Kinetic analysis of S. cerevisiae AK revealed K(m) values of (3.5+/-0.2) micromol/L for adenosine and (100.0+/-11.0) micromol/L for ATP, with k(cat) of (1530+/-20) min(-1) for adenosine and (1448+/-25) min(-1) for ATP. The determination of the K(m) value for other nucleosides and deoxynucleoside indicated that the nucleoside specificity of this enzyme from yeast was quite high.