title: Physiological role of FolD (methylenetetrahydrofolate dehydrogenase), FchA (methenyltetrahydrofolate cyclohydrolase) and Fhs (formyltetrahydrofolate synthetase) from Clostridium perfringens in a heterologous model of Escherichia coli.
abstract: Most organisms possess bifunctional FolD [5,10-methylenetetrahydrofolate (5,10-CH2-THF) dehydrogenase-cyclohydrolase] to generate NADPH and 10-formyltetrahdrofolate (10-CHO-THF) required in various metabolic steps. In addition, some organisms including Clostridium perfringens possess another protein, Fhs (formyltetrahydrofolate synthetase), to synthesize 10-CHO-THF. Here, we show that unlike the bifunctional FolD of Escherichia coli (EcoFolD), and contrary to its annotated bifunctional nature, C. perfringens FolD (CpeFolD) is a monofunctional 5,10-CH2-THF dehydrogenase. The dehydrogenase activity of CpeFolD is about five times more efficient than that of EcoFolD. The 5,10-methenyltetrahydrofolate (5,10-CH+-THF) cyclohydrolase activity in C. perfringens is provided by another protein, FchA (5,10-CH+-THF cyclohydrolase), whose cyclohydrolase activity is ∼ 10 times more efficient than that of EcoFolD. Kinetic parameters for CpeFhs were also determined for utilization of all of its substrates. Both CpeFolD and CpeFchA are required to substitute for the single bifunctional FolD in E. coli. The simultaneous presence of CpeFolD and CpeFchA is also necessary to rescue an E. coli folD deletion strain (harbouring CpeFhs support) for its formate and glycine auxotrophies, and to alleviate its susceptibility to trimethoprim (an antifolate drug) or UV light. The presence of the three clostridial proteins (FolD, FchA and Fhs) is required to maintain folate homeostasis in the cell.
authors.affiliation: Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Srinivas
authors.initials: S
authors.lastname: Aluri
authors.affiliation: Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Shivjee
authors.initials: S
authors.lastname: Sah
authors.affiliation: Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Sandeep
authors.initials: S
authors.lastname: Miryala
authors.affiliation: Jawaharlal Nehru Centre for Advanced Scientific Research, Bangalore 560064, India.|Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Umesh
authors.initials: U
authors.lastname: Varshney
mesh_terms: D000595:Amino Acid Sequence; D003016:Clostridium perfringens / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D004926:Escherichia coli / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D005574:Formate-Tetrahydrofolate Ligase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D045064:Methenyltetrahydrofolate Cyclohydrolase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008754:Methylenetetrahydrofolate Dehydrogenase (NADP) / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016415:Sequence Alignment
chemical_list: D008754:Methylenetetrahydrofolate Dehydrogenase (NADP); D045064:Methenyltetrahydrofolate Cyclohydrolase; D005574:Formate-Tetrahydrofolate Ligase