Isolation and sequencing of the cDNA coding for spinach 10-formyltetrahydrofolate synthetase. Comparisons with the yeast, mammalian, and bacterial proteins.
The one-carbon metabolism enzymes 10-formyltetrahydrofolate synthetase (EC 6.3.4.3), 5,10-methenyltetrahydrofolate cyclohydrolase (EC 3.5.4.9), and 5,10-methylenetetrahydrofolate dehydrogenase (EC 1.5.1.5) can be found on a single trifunctional protein in the eukaryotes examined. The one exception is in spinach leaves where 10-formyltetrahydrofolate synthetase is monofunctional (Nour, J. M., and Rabinowitz, J. C. (1991) J. Biol. Chem. 266, 18363-18369). In the prokaryotes examined, 10-formyltetrahydrofolate synthetase is either absent or is monofunctional. A cDNA clone encoding spinach leaf 10-formyltetrahydrofolate synthetase was isolated through the use of antibodies to the purified enzyme. This clone had an open reading frame of 1914 base pairs and encoded for a protein containing 636 amino acids with a calculated M(r) of 67,727. The percentage identity between spinach 10-formyltetrahydrofolate synthetase and the synthetase domains in the four trifunctional eukaryotic enzymes and the two monofunctional prokaryotic enzymes that have been cloned and sequenced was: 64.9% human, 63.8% rat, 55.6% yeast cytoplasm, 53.8% yeast mitochondria, 47.8% Clostridium acidi-urici, and 47.9% Clostridium thermoaceticum. Clearly the spinach monofunctional protein had greatest homology with the mammalian proteins. The spinach protein is longer than the two other monofunctional prokaryotic proteins. Possible reasons for this are presented. The codon usage and the putative translation initiation sites are examined and compared with other spinach proteins.
Nour
J M
JM
Department of Molecular and Cell Biology, University of California, Berkeley 94720.
Rabinowitz
J C
JC
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D003013:Clostridium / Q000201:enzymology / Q000235:genetics*; D004247:DNA / Q000235:genetics / Q000302:isolation & purification*; D005574:Formate-Tetrahydrofolate Ligase / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010944:Plants / Q000201:enzymology* / Q000235:genetics*; D051381:Rats; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D012689:Sequence Homology, Nucleic Acid
D004247:DNA; D005574:Formate-Tetrahydrofolate Ligase