PURIFICATION AND CHARACTERIZATION OF POLYPHOSPHATASE FROM SACCHAROMYCES-CEREVISIAE CYTOSOL

A homogenous polyphosphatase preparation was isolated from Saccharomyc es cerevisiae cytosol. The specific activity of the enzyme was 283 U/m g protein, and it was obtained with 3.8% yield and 3540-fold purificat ion. The polyphosphatase is monomeric with molecular mass similar to 4 0 kD. The enzyme hydrolyzes polyphosphates with various chain lengths, including tripolyphosphate; it is essentially inactive with ATP, PPi, and p-nitrophenyl phosphate. The enzyme is most active at 50 degrees C and pH 6.5-8.5. The enzyme activity increases 8-66-fold in the prese nce of some divalent cations, with the degree of stimulation being in the order: Co2+< Mn2+ > Mg2+ > Zn2+ > Fe2+. The polyphosphatase is ina ctive with Ca2+ and Cu2+. Heparin and antibodies against purified cell envelope polyphosphatase as well as Cu2+ and Zn2+ in the presence of Mg2+ are effective inhibitors of the cytosol polyphosphatase. The cyto sol polyphosphatase is quite similar to purified cell envelope polypho sphatase and differs in a number of properties from polyphosphatases o f vacuoles, nuclei, and mitochondria isolated from the same yeast.