SEA-PHAGES | thymidylate synthases versus dihydrofolate reductases

Link to this post \| posted 29 May, 2018 17:34
smolloy123	Hi All, I am finishing up a Microbacterium phage annotation, cluster EB phage Eden. Homologues of Eden gp57 are all called as thymidylate synthases…why? There are strong HHPRED results for a dihydrofolate reductase (see attachment or Pecaan site for Eden gene stopping at 36,681. BlastP at NCBI also points towards dihdrofolate reductases in bacterial species. Does anyone know why these genes are called as thymidylate synthases in the other EB phage? Do I go with the phamerator/phasgedb flow on this or go with the evidence? I thought maybe there is some other evidence that I don't know about that points towards thymidylate synthase. This just doesn't look like the latter to me. Thanks! Sally 344Kb

Link to this post | posted 29 May, 2018 17:34

Hi All,
I am finishing up a Microbacterium phage annotation, cluster EB phage Eden. Homologues of Eden gp57 are all called as thymidylate synthases…why? There are strong HHPRED results for a dihydrofolate reductase (see attachment or Pecaan site for Eden gene stopping at 36,681. BlastP at NCBI also points towards dihdrofolate reductases in bacterial species. Does anyone know why these genes are called as thymidylate synthases in the other EB phage? Do I go with the phamerator/phasgedb flow on this or go with the evidence? I thought maybe there is some other evidence that I don't know about that points towards thymidylate synthase. This just doesn't look like the latter to me.
Thanks!
Sally

Link to this post \| posted 29 May, 2018 21:26
cdshaffer	I think this is mostly an issue with synonyms and specificity of functional names. A quick check of Kegg shows that there are three entries for "thymidylate synthases". K00560 thyA, TYMS; thymidylate synthase [EC:2.1.1.45] K03465 thyX, thy1; thymidylate synthase (FAD) [EC:2.1.1.148] K13998 DHFR-TS; dihydrofolate reductase / thymidylate synthase [EC:1.5.1.3 2.1.1.45] Notice how the first and the last share a common EC number (ie they can carry out the same reaction). If you dig deeper into these entries you will see that all the thymidylate synthases are methytransferases and all three use dihydrofolate as the source of the methyl group and convert dUMP to dTMP (i.e. they are both a dihydofolate reductase and a thymidylate synthase). If you look farther down on the HHPRED search you will also see entries like this that are only ever so slightly lower probabilities which is also a hint as to the nature of the issue: 1J3K_A Bifunctional dihydrofolate reductase (E.C.1.5.1.3)-thymidylate synthase; bifunctional, OXIDOREDUCTASE, TRANSFERASE; HET: NDP, UMP, WRA; 2.1A {Plasmodium falciparum} SCOP: c.71.1.1 As for annotation, the current approved term is "ThyX-like thymidylate synthase"; but I am not sure that should be used here. Based on my brief investigation it appears that ThyX uses FADH while ThyA/DHFR-TS use NADH as coenzymes (hence the different EC numbers). Since all your HHPRED hits are to the DHFR-TS proteins they are likely not really "ThyX type" but rather a "ThyA type". So then the proper term used here should maybe be "ThyA-like thymidylate synthase" or simply "thymidylate synthase"; but someone should really double check the enzymology before adding anything to the approved list. Edited 29 May, 2018 21:30

Link to this post | posted 29 May, 2018 21:26

cdshaffer

I think this is mostly an issue with synonyms and specificity of functional names. A quick check of Kegg shows that there are three entries for "thymidylate synthases".

K00560
thyA, TYMS; thymidylate synthase [EC:2.1.1.45]
K03465
thyX, thy1; thymidylate synthase (FAD) [EC:2.1.1.148]
K13998
DHFR-TS; dihydrofolate reductase / thymidylate synthase [EC:1.5.1.3 2.1.1.45]

Notice how the first and the last share a common EC number (ie they can carry out the same reaction). If you dig deeper into these entries you will see that all the thymidylate synthases are methytransferases and all three use dihydrofolate as the source of the methyl group and convert dUMP to dTMP (i.e. they are both a dihydofolate reductase and a thymidylate synthase). If you look farther down on the HHPRED search you will also see entries like this that are only ever so slightly lower probabilities which is also a hint as to the nature of the issue:

1J3K_A Bifunctional dihydrofolate reductase (E.C.1.5.1.3)-thymidylate synthase; bifunctional, OXIDOREDUCTASE, TRANSFERASE; HET: NDP, UMP, WRA; 2.1A {Plasmodium falciparum} SCOP: c.71.1.1

As for annotation, the current approved term is "ThyX-like thymidylate synthase"; but I am not sure that should be used here. Based on my brief investigation it appears that ThyX uses FADH while ThyA/DHFR-TS use NADH as coenzymes (hence the different EC numbers). Since all your HHPRED hits are to the DHFR-TS proteins they are likely not really "ThyX type" but rather a "ThyA type". So then the proper term used here should maybe be "ThyA-like thymidylate synthase" or simply "thymidylate synthase"; but someone should really double check the enzymology before adding anything to the approved list.

Edited 29 May, 2018 21:30

Link to this post \| posted 30 May, 2018 00:52
debbie	I think that when I called this, I decided that thymidylate synthase is a generic enough term that it would be best until more is known. I unfortunately didn't put the more generic term "thymidylate synthase" on the approved list (until today). I didn't use dihydrofolate reductase because it wasn't on the list; it surely could be. Neither are very specific or satisfying, using both seems to be too much. We could add ThyA-like thymidylate synthase, but until someone wants to look at all thymidylate synthases more closely, I think it could be problematic. Chris - thank you for your investigation on this, it was very helpful.

Link to this post | posted 30 May, 2018 00:52

debbie

I think that when I called this, I decided that thymidylate synthase is a generic enough term that it would be best until more is known. I unfortunately didn't put the more generic term "thymidylate synthase" on the approved list (until today). I didn't use dihydrofolate reductase because it wasn't on the list; it surely could be. Neither are very specific or satisfying, using both seems to be too much. We could add ThyA-like thymidylate synthase, but until someone wants to look at all thymidylate synthases more closely, I think it could be problematic. Chris - thank you for your investigation on this, it was very helpful.

Link to this post \| posted 31 May, 2018 12:36
smolloy123	Hi Chris, Thanks for this. Super helpful!

Link to this post \| posted 31 May, 2018 12:37
smolloy123	Debbie Jacobs-Sera I think that when I called this, I decided that thymidylate synthase is a generic enough term that it would be best until more is known. I unfortunately didn't put the more generic term "thymidylate synthase" on the approved list (until today). I didn't use dihydrofolate reductase because it wasn't on the list; it surely could be. Neither are very specific or satisfying, using both seems to be too much. We could add ThyA-like thymidylate synthase, but until someone wants to look at all thymidylate synthases more closely, I think it could be problematic. Chris - thank you for your investigation on this, it was very helpful. Hi Deb, I'll go with he generic thymidylate synthase. Thanks to both you and Chris for helping me sort this cheers Sally

Link to this post | posted 31 May, 2018 12:37

smolloy123

Debbie Jacobs-Sera
I think that when I called this, I decided that thymidylate synthase is a generic enough term that it would be best until more is known. I unfortunately didn't put the more generic term "thymidylate synthase" on the approved list (until today). I didn't use dihydrofolate reductase because it wasn't on the list; it surely could be. Neither are very specific or satisfying, using both seems to be too much. We could add ThyA-like thymidylate synthase, but until someone wants to look at all thymidylate synthases more closely, I think it could be problematic. Chris - thank you for your investigation on this, it was very helpful.

Hi Deb,
I'll go with he generic thymidylate synthase. Thanks to both you and Chris for helping me sort this smile

cheers
Sally

Link to this post \| posted 26 Apr, 2019 16:50
dbolliva	I have been looking at the genes in Celaena, our new EB phage and see two genes close to each other (stops 38540 and 39809) that both were giving HHPred results in the 99-100 % probablility range for the bifunctional DHFR-TS. I went to look at the PDB files for the 1J3K structure and noticed that the two functions are in separate domains. The A and B chains carry the DHFR activity and the and D chains have the thymidylates synthase domains. Gene 55 of Celaena (stop 38540) is aligning with the A and B chains and Gene 57 of Celaena (stop 39809) aligns with the C and D chains. Based on this I was wondering if all of the pham members of gene 55 should be DHFR (like Bubbabear_56) and all of the pham members of gene 57 should be labeled thymidylate synthase like BubbaBear_59?

Link to this post | posted 26 Apr, 2019 16:50

dbolliva

I have been looking at the genes in Celaena, our new EB phage and see two genes close to each other (stops 38540 and 39809) that both were giving HHPred results in the 99-100 % probablility range for the bifunctional DHFR-TS. I went to look at the PDB files for the 1J3K structure and noticed that the two functions are in separate domains. The A and B chains carry the DHFR activity and the and D chains have the thymidylates synthase domains. Gene 55 of Celaena (stop 38540) is aligning with the A and B chains and Gene 57 of Celaena (stop 39809) aligns with the C and D chains. Based on this I was wondering if all of the pham members of gene 55 should be DHFR (like Bubbabear_56) and all of the pham members of gene 57 should be labeled thymidylate synthase like BubbaBear_59?

Link to this post \| posted 26 Apr, 2019 18:32
welkin	Hi Dave, Sounds like it to me. Thanks!

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thymidylate synthases versus dihydrofolate reductases