Subsystem: 5-FCL-like protein
This subsystem's description is:
HYPOTHESIS: In spite of high homology with bona fide 5-Formyltetrahydrofolate cycloligases (5-FCL), the 5-FCL-like protein family (which includes At1g76730 and its homologs in plans, animal, archaea, and bacteria) is predicted not to have 5-formyltetrahydrofolate cycloligase activity. Potentially involved in involved in thiamine salvage
1. 5-Formyltetrahydrofolate (5-CHO-THF) is an inhibitory folate formed from 5,10-methenyltetrahydrofolate (5,10-CH=THF) in a side-reaction mediated by serine hydroxymethyltransferase (GlyA). 5-Formyltetrahydrofolate cycloligase (5-FCL, also called methenyltetrahydrofolate synthetase) is an ATP-dependent enzyme that reconverts 5-CHO-THF to 5,10-CH=THF. 5-FCL, like GlyA, occurs in most bacteria and is sometimes clustered with GlyA or other genes of folate-dependent C1 metabolism. 5-FCL also occurs in all eukaryotes. Basically, 5-FCL is needed in organisms with GlyA to ‘detoxify’ 5-CHO-THF.
2. A protein weakly similar to 5-FCL (annotated in SEED as ‘5-FCL-like protein’ occurs in plants, animals, many archaea, and occasional bacteria. It belongs to COG0212 (as do true 5-FCL enzymes). The BlastP scores are as high as e-50 between the 5-FCL-like proteins from these groups. For comparison, the BlastP scores between 5-FCL-like proteins and true 5-FCL proteins are typically e-03 or poorer.
3. 3D structure prediction programs (COMPASS, PHYRE, GenThreader) all give highly significant matches between 5-FCL-like proteins from plants, animals, or archaea and canonical 5-FCL structures.
4. Some vertebrate 5-FCL-like proteins are fused to a C-terminal RNA recognition motif. There are no other informative fusions in eukaryotes or prokaryotes.
5. Some association evidence links 5-FCL-like genes to folates or C1 metabolism:
- In Aeropyrum pernix the 5-FCL-like gene is translationally coupled to a MetF family protein.
- In Halomicrobium mukohatei it is adjacent to a probable formate transporter
- In Synechococcus PCC 7002 it is next-but-one to GlyA.
- In Bacillus halodurans it is adjacent to a SAM riboswitch (on the opposite strand).
6. Some bioinformatic evidence suggests that 5-FCL-like proteins do not have 5-FCL activity:
- Where it occurs outside Archaea (bacteria, plants, animals) there is almost always also a canonical 5-FCL in the genome (exception: Syntrophobacter fumaroxidans)
- It occurs in some archaea (e.g., Sulfobolus, Methanothermobacter) without folD, puN, purH, i.e. with no way to metabolize 5,10-CH=THF and most probably little if any 5,10-CH=THF substrate for SHMT-mediated 5-formyl-THF formation.
- It occurs in some archaea (e.g., Sulfobolus, Methanothermobacter) that have a SHMT that uses modified folates, not THF derivatives, and which therefore presumably cannot form 5-formyl-THF.
- Along with 5-FCL, it is absent from some archaea that have folates, e.g. Haloquadratum, Halorhabdus, Methanosarcina barkeri).
- The presumptive 5-FCL active site is conserved but the region upstream of this site differs greatly.
7. Summary of experimental evidence on 5-FCL-like genes:
- The Arabidopsis 5-FCL-like gene is essential; the Arabidopsis 5-FCL-like protein fused to GFP is targeted to plastids.
- The Haloferax 5-FCL-like gene is not essential; the knockout grows normally. Analysis of folate pools shows no consistent differences from wild type. If it were a 5-FCL, accumulation of 5-CHO-THF would be expected. (This is observed in E. coli when the canonical 5-FCL YgfA is deleted.)
- Wild type Haloferax does not assimilate 14C-formate, so that the 5-FCL-like protein cannot be a novel 10-formyltetrahydrofolate synthetase.
- The recombinant 5-FCL-like proteins of Bacillus halodurans and Arabidopsis do not have detectable 5-FCL activity in vitro.
- The Synechococcus PCC7002 5-FCL-like gene failed to complement the E. coli 5FCL (ygfA) mutant (which cannot grow on glycine as sole N source). The Bacillus halodurans 5-FCL-like gene may complement the ygfA mutant weakly but the complemented colonies cannot be propagated; more generally, the B. halodurans 5-FCL-like gene is toxic to E. coli grown on minimal medium.
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| Diagram | Functional Roles | Subsystem Spreadsheet | Description | |||||||||
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HYPOTHESIS: In spite of high homology with bona fide 5-Formyltetrahydrofolate cycloligases (5-FCL), the 5-FCL-like protein family (which includes At1g76730 and its homologs in plans, animal, archaea, and bacteria) is predicted not to have 5-formyltetrahydrofolate cycloligase activity. Potentially involved in involved in thiamine salvage 1. 5-Formyltetrahydrofolate (5-CHO-THF) is an inhibitory folate formed from 5,10-methenyltetrahydrofolate (5,10-CH=THF) in a side-reaction mediated by serine hydroxymethyltransferase (GlyA). 5-Formyltetrahydrofolate cycloligase (5-FCL, also called methenyltetrahydrofolate synthetase) is an ATP-dependent enzyme that reconverts 5-CHO-THF to 5,10-CH=THF. 5-FCL, like GlyA, occurs in most bacteria and is sometimes clustered with GlyA or other genes of folate-dependent C1 metabolism. 5-FCL also occurs in all eukaryotes. Basically, 5-FCL is needed in organisms with GlyA to ‘detoxify’ 5-CHO-THF. 2. A protein weakly similar to 5-FCL (annotated in SEED as ‘5-FCL-like protein’ occurs in plants, animals, many archaea, and occasional bacteria. It belongs to COG0212 (as do true 5-FCL enzymes). The BlastP scores are as high as e-50 between the 5-FCL-like proteins from these groups. For comparison, the BlastP scores between 5-FCL-like proteins and true 5-FCL proteins are typically e-03 or poorer. 3. 3D structure prediction programs (COMPASS, PHYRE, GenThreader) all give highly significant matches between 5-FCL-like proteins from plants, animals, or archaea and canonical 5-FCL structures. 4. Some vertebrate 5-FCL-like proteins are fused to a C-terminal RNA recognition motif. There are no other informative fusions in eukaryotes or prokaryotes. 5. Some association evidence links 5-FCL-like genes to folates or C1 metabolism: - In Aeropyrum pernix the 5-FCL-like gene is translationally coupled to a MetF family protein. - In Halomicrobium mukohatei it is adjacent to a probable formate transporter - In Synechococcus PCC 7002 it is next-but-one to GlyA. - In Bacillus halodurans it is adjacent to a SAM riboswitch (on the opposite strand). 6. Some bioinformatic evidence suggests that 5-FCL-like proteins do not have 5-FCL activity: - Where it occurs outside Archaea (bacteria, plants, animals) there is almost always also a canonical 5-FCL in the genome (exception: Syntrophobacter fumaroxidans) - It occurs in some archaea (e.g., Sulfobolus, Methanothermobacter) without folD, puN, purH, i.e. with no way to metabolize 5,10-CH=THF and most probably little if any 5,10-CH=THF substrate for SHMT-mediated 5-formyl-THF formation. - It occurs in some archaea (e.g., Sulfobolus, Methanothermobacter) that have a SHMT that uses modified folates, not THF derivatives, and which therefore presumably cannot form 5-formyl-THF. - Along with 5-FCL, it is absent from some archaea that have folates, e.g. Haloquadratum, Halorhabdus, Methanosarcina barkeri). - The presumptive 5-FCL active site is conserved but the region upstream of this site differs greatly. 7. Summary of experimental evidence on 5-FCL-like genes: - The Arabidopsis 5-FCL-like gene is essential; the Arabidopsis 5-FCL-like protein fused to GFP is targeted to plastids. - The Haloferax 5-FCL-like gene is not essential; the knockout grows normally. Analysis of folate pools shows no consistent differences from wild type. If it were a 5-FCL, accumulation of 5-CHO-THF would be expected. (This is observed in E. coli when the canonical 5-FCL YgfA is deleted.) - Wild type Haloferax does not assimilate 14C-formate, so that the 5-FCL-like protein cannot be a novel 10-formyltetrahydrofolate synthetase. - The recombinant 5-FCL-like proteins of Bacillus halodurans and Arabidopsis do not have detectable 5-FCL activity in vitro. - The Synechococcus PCC7002 5-FCL-like gene failed to complement the E. coli 5FCL (ygfA) mutant (which cannot grow on glycine as sole N source). The Bacillus halodurans 5-FCL-like gene may complement the ygfA mutant weakly but the complemented colonies cannot be propagated; more generally, the B. halodurans 5-FCL-like gene is toxic to E. coli grown on minimal medium. | ||||||||||||
