232 Gray and LuThe BAL 31 nucleases are remarkable for their resistance to inac-
tivation and or denaturation in the presence of detergents, urea, or high
concentrations of electrolyte, and are very stable upon extended stor-
age in the cold (3,4, 8). The nuclease activities are not highly resistant
to inactivation at elevated temperatures, but the overall secondary and
tertiary structures of the S enzyme are extremely resistant to disrup-
tion: The internally proteolytically cleaved enzyme fails to dissociate
at 100°C in the presence of 1% sodium dodecyl sulfate (7).
1.3. Reactions Catalyzed
The nucleases have three general activities against DNA: a 3'-5'
exonuclease activity that apparently removes one residue at a time
from duplex structures (9), a 5'-3' exonuclease activity that carries out
the bulk of the degradation of single-stranded DNA (10), and a much
slower (2-3% of the bond cleavage rate for the 5'-3' exonuclease [10])
endonuclease activity against single-stranded DNA that is also elic-
ited by a variety of covalent and noncovalent lesions or distortions in
duplex DNA (3,4, 6,8,11-14). The two forms differ greatly in the rates
of catalysis, at given molar concentrations of duplex ends and enzyme,
for the exonuclease reaction if the DNA is double-stranded (hence the
aforementioned "fast" and "slow" designations); but the two forms are
much more comparable in kinetic behavior if the substrate is single-
stranded DNA (6,10). The nucleases are not sugar-specific and cata-
lyze the terminally directed hydrolysis of duplex RNA and readily
degrade RNA containing nonduplex structure (15). Endonucleolytic
cleavage of duplex RNA in response to lesions or distortions has not
been examined except that it is likely that the nucleases can cleave in
response to a strand break in duplex RNA (15).
The combination of a 3'-5' exonuclease activity on duplex DNA, a
5'-3' mode of attack on single-stranded DNA, and relatively infrequent
endonucleolytic attack in single-stranded DNA serves to reduce the
length of linear duplex DNA and is referred to as the duplex exonu-
clease activity. Partially degraded molecules possess the expected 5'-
terminated single-stranded "tails," and there was no evidence for
3'-terminated tails (9), implying that the 5'-3' attack is limited to single-
stranded DNA. An unexpected aspect of the mechanism is that the
average length of the tails, for a constant number of nucleotides removed
by the exonuclease action, decreases markedly to a limiting value with