The exudate of fully germinated spores of Bacillus cereus IF0 13597 in 0.25 M sodium phosphate buffer, pH 7-08 was found to contain a spore-lytic enzyme. This enzyme was found to cause loss of absorbance in coat-stripped spore suspensions and phase-darkening of the spores but had minimal activity on isolated peptidoglycan substrates. The enzyme was purified in an active form and identified as a 24 kDa protein which is either an amidase or a peptidase. The amino-terminal 19 residues had the following sequence: FSNQVIQRGASGEKVIELQ. The spore-lytic enzyme retained its activity in a medium of a relatively high ionic strength containing a non-ionic surfactant such as nonaethyleneglycol n-dodecyl ether. This activity was optimum at a salt concentration of about 30 mM in assay buffer at neutral pH. In contrast to the enzyme in a spore-bound form, the enzyme in solution was shown to be heat-sensitive and was readily inactivated by thiol reagents.
ISG20 is an interferon-induced antiviral exoribonuclease that acts on single-stranded RNA and also has minor activity towards single-stranded DNA. It belongs to the DEDDh group of RNases of the DEDD exonuclease superfamily. We have solved the crystal structure of human ISG20 complexed with two Mn 2þ ions and uridine 5 0 -monophosphate (UMP) at 1.9 A resolution. Its structure, including that of the active site, is very similar to those of the corresponding domains of two DEDDh-group DNases, the e subunit of Escherichia coli DNA polymerase III and E. coli exonuclease I, strongly suggesting that its catalytic mechanism is identical to that of the two DNases. However, ISG20 also has distinctive residues, Met14 and Arg53, to accommodate hydrogen bonds with the 2 0 -OH group of the UMP ribose, and these residues may be responsible for the preference of ISG20 for RNA substrates.
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