THE PARTIAL-PURIFICATION AND CHARACTERIZATION OF PURINE NUCLEOSIDE PHOSPHORYLASE FROM MAMMALIAN MITOCHONDRIA

Cytosolic purine nucleoside phosphorylase (PNPase) is a well known, an d described enzyme which exists in a variety of organisms, both procar yotic and eucaryotic. More recently this enzyme was found in bovine li ver mitochondria. The mitochondrial purine nucleoside phosphorylase wa s purified 63 fold and has a molecular weight of 48-60 kD. From Linewe aver-Burk plots apparent K-M's of 23 mu M for inosine, 42 mu M for deo xyinosine, 40 mu M for phosphate, 2 mu M for hypoxanthine, and 163 mu M for ribose-1-phosphate were calculated. Both 8-aminoguanosine (K-i = 0.5 mu M) and araG (K-i = 381 mu M) are inhibitors of the enzyme. The protein's isoelectric point (pI) was calculated at a pH of 4.2. Preli minary immunological work showed no cross-reactivity between epitopes on the mitochondrial protein and those on PNPase from human erythrocyt es. The apparent K-M's calculated for the mitochondrial enzyme are, wi th the exception of that using hypoxanthine, within the range commonly associated with K-M's from the cytosolic species. The mitochondrial e nzyme's molecular weight and pI are less than normally described. The enzyme's isolation from mitochondria, together with several unique cha racteristics, suggest that it is a separate protein from that found in the cytosol.