THE humAL OF BroLocrcAL CHE.%mTRY

Exonuclease activity other than RNase II (EC 3.1.4.23) was detected and purified from Escherichia coli, using [“HlrRNA, T4 phage-specific mRNA, and poly(U) as substrates. The activity co-sedimented with ribosomes and tended to aggregate in 10 rnM Tris/Cl (pH 7.51, 10 mM MgC12, but sedimented free of ribosomes and was more stable in buffer containing 1 M KCl. It was purified 300-fold by chromatography on DEAE-cellulose and hydroxyapatite or phosphocellulose. From substrates labeled uniformly with L3Hluridine, and at their 5’ termini with ?lP, the activity, like RNase II, preferentially released “H label. The activity was distinguishable from RNase II in that it: (a) bound tightly to hydroxyapatite and phosphocellulose; (b) eluted from Sephadex G-200 with one-half to two-thirds the apparent size of RNase II; (c) was present in extracts deficient in RNase II, and was unchanged in its heat stability in a strain with thermolabile RNase II; (d) degraded rRNA at least as well as it did poly(U); and (e) aggregated in low ionic strength buffers. The activity eluted from phosphocellulose columns in three major peaks. These seem to be forms of a single new enzyme since: (a) all of them released 5’-XMP without a lag as the only alcoholor acid-soluble product of RNA; (b) all showed maximal activity at 0.1 to 0.2 M KC1 or NH&l and 0.1 mM MgCl,; and (c) all were reduced more than 20fold in two independently isolated mutant strains.