J. Biol. Chem., Vol. 268, Issue 32, 23792-23798, Nov, 1993
Single-stranded DNA binding activity of C1-tetrahydrofolate synthase enzymes
WP Wahls, JM Song and GR Smith
Fred Hutchinson Cancer Research Center, Seattle, Washington 98104.
In eukaryotes C1-5,6,7,8-tetrahydrofolate (THF) synthase is a trifunctional
enzyme that catalyzes the interconversion of reduced forms of folate to
supply activated one-carbon units required for a variety of metabolic
pathways. The enzymatic activities include 10- formyl-THF synthetase (EC
6.3.4.3), 5,10-methenyl-THF cyclohydrolase (EC 3.5.4.9), and
5,10-methylene-THF dehydrogenase (EC 1.5.1.5). In bacteria separate,
monofunctional or bifunctional polypeptides catalyze the same reactions. We
have purified C1-THF synthase from the fission yeast Schizosaccharomyces
pombe and found its physical and enzymatic properties similar to those of
other eukaryotic C1-THF synthase enzymes. Unexpectedly, the S. pombe enzyme
bound strongly (Keq = 100 pM) to single-stranded DNA, but not to
double-stranded DNA or to RNA. The binding was sequence-independent,
apparently not cooperative, and not detectably inhibited by C1-THF synthase
substrates or cofactors. Trifunctional cytoplasmic enzyme from
Saccharomyces cerevisiae and monofunctional (synthetase) enzyme from
Clostridium acidiurici also bound tightly to single-stranded DNA, while
bifunctional (dehydrogenase and cyclohydrolase) enzyme from Escherichia
coli did not, suggesting that single-stranded DNA binding is a conserved
function of the synthetase domain of C1-THF synthase enzymes.
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