Physical map location of the Escherichia coli gene encoding the bifunctional enzyme 5,10-methylene-tetrahydrofolate dehydrogenase/5,10-methenyl-tetrahydrofolate cyclohydrolase

Oxidation of 5,10-methylene-tetrahydrofolate is catalyzed by methylene-tetrahydrofolate dehydrogenase (EC 1.5.1.5): The enzyme was partially purified from Escherichia coli (2, 3) and has now been purified to homogeneity from that source (1). It was concluded that the E. coli enzyme is a bifunctional enzyme possessing, in addition to dehydrogenase activity, 5,10-methenyl-tetrahydrofolate cyclohydro- lase (EC 3.5.4.9) activity (1, 2): [6R]-5,10-methenyl+-THF + H20 = [6R]-10-formyl-THF + HW. The gene coding for the enzyme has been cloned and sequenced (1), and its position on the physical map of the E. coli chromosome was determined by using a gene mapping membrane kindly provided by Akihiro Noda. The membrane contained the 476-clone miniset that includes the entire E. coli genome (4). The probe was a 25-mer oligonucleotide that exactly matches the sequence coding for amino acids 9 to 17 of the amino-terminal region of the enzyme. The positive clones were 157 and 158.