#11552 query GO:0052928 CTP:3'-cytidine-tRNA cytidylyltransferase activity

[Valerie Wood]

I am trying to figure the differenece between these:

GO:0052928 CTP:3'-cytidine-tRNA cytidylyltransferase activity
Catalysis of the reaction: a tRNA with a 3' cytidine + CTP = a tRNA with a 3' CC end + diphosphate.

GO:0052927 CTP:tRNA cytidylyltransferase activity
Catalysis of the reaction: a tRNA precursor + CTP = a tRNA with a 3' cytidine end + diphosphate.

the genes i am annotating seem to be annotated to both and they have identical EC number.
What is a CC end?

val

[David Osumi-Sutherland]

Seems to come directly from KEGG

http://www.genome.jp/dbget-bin/www_bget?rn:R09384
http://www.genome.jp/dbget-bin/www_bget?rn:R09383

[Peter D\'Eustachio]

The GONUTS page for GO:0052928 (http://gowiki.tamu.edu/wiki/index.php/Category:GO:0052928_!_CTP:3%27-cytidine-tRNA_cytidylyltransferase_activity) and its ExPASY ENZYME link look like they might be helpful here, hinting at a description of CC and also at a possible history of EC term merger (so the two GO terms could possibly point to two activities that are now regarded as one by the enzymologists).

[David Osumi-Sutherland]

Interesting that the EC term def at http://enzyme.expasy.org/EC/2.7.7.72 has a different reaction to either of the above:

A tRNA precursor + 2 CTP + ATP <=> a tRNA with a 3' CCA end + 3 diphosphate

This page also has a nice description:
The CCA sequence is added to the tRNA precursor by stepwise nucleotide addition performed by a single enzyme that is ubiquitous in all living organisms. Although the enzyme has the option of releasing the product after each addition, it prefers to stay bound to the product and proceed with the next addition.

It also mentions EC mergers: Formerly EC 2.7.7.21 and EC 2.7.7.25.

Tracing original IDs:
http://www.genome.jp/dbget-bin/www_bget?ec:2.7.7.25
http://www.genome.jp/dbget-bin/www_bget?ec:2.7.7.21

The old names look consistent with the original entries being for separate enzymes adding CC and adding A, being merged to one activity which adds CCA.

This KEGG page seems to allow for both substrates: http://www.genome.jp/dbget-bin/www_bget?ec:2.7.7.72 :

Substrate
tRNA precursor [CPD:C02211];
CTP [CPD:C00063];
ATP [CPD:C00002];
tRNA with a 3' cytidine [CPD:C19078];
tRNA with a 3' CC end [CPD:C19080]
Product
tRNA with a 3' CCA end [CPD:C19085];
diphosphate [CPD:C00013];
tRNA with a 3' cytidine end;
tRNA with a 3' CC end [CPD:C19080]

This is consistent with tRNA with a 3' CC end [CPD:C19080] as an intermediate that is sometimes release.

So, I think you're right that these should be merged.

[David Osumi-Sutherland]

Looks like these two should be merged too:

id "GO:0004810"^^string
label "tRNA adenylyltransferase activity"^^string
comment "This term represents a deleted EC activity and is scheduled for obsoletion."^^string
database_cross_reference "EC:2.7.7.25"^^string

id "GO:0052929"^^string
label "ATP:3'-cytidine-cytidine-tRNA adenylyltransferase activity"^^string
database_cross_reference "EC:2.7.7.72"^^string
database_cross_reference "KEGG:R09386"^^string
definition "Catalysis of the reaction: a tRNA with a 3' CC end + ATP = a tRNA with a 3' CCA end + diphosphate."^^string

I guess it doesn't really matter which term wins in the merge so I'll chose the one with most annotations to reduce churn. The new term will take its definition from the Expasy page. Does that seem reasonable?

[Valerie Wood]

Sounds reasonable to me. See what Peter thinks.

val

[Peter D\'Eustachio]

Sounds reasonable to me too.
Peter

[Harold J. Drabkin]

One might want to look at:PMID:18523015
Evolution of tRNA nucleotidyltransferases: a small deletion generated CC-adding enzymes.
" In some organisms, CCA synthesis is accomplished by the collaboration of evolutionary closely related enzymes with partial activities (CC and A addition). These enzymes carry all known motifs of the catalytic core found in CCA-adding enzymes. Therefore, it is a mystery why these polymerases are restricted in their activity and do not synthesize a complete CCA terminus. "

In some organisms ONE enzyme can add the whole CCA; In some bacteria, however, it appears TWO enzymes, one to add CC, a separate one to add the final A.
Also note that in eukaryotes, CCA is not encoded and always has to be added after removal of 3' trailer, but in bacteria, CCA is encoded; 3' end maturation stops at the CCA.
However, bacteria still do have CCA adding enzymes to maintain the CCA

[Peter D\'Eustachio]

Hi Harold,
Okay, in different reactions different units get added to the 3' end of the tRNA. But if it's mechanistically the same reaction, then couldn't a single GO molecular function term could cover all the reactions?
Peter

[David Osumi-Sutherland]

Thanks Harold. So I guess we need 3 terms. Do these look OK?

CCA tRNA nucleotidyltransferase activity (<- new?)
A tRNA precursor + 2 CTP + ATP <=> a tRNA with a 3' CCA end + 3 diphosphate

ATP:3'-cytidine-cytidine-tRNA adenylyltransferase activity
Catalysis of the reaction: a tRNA with a 3' CC end + ATP = a tRNA with a 3' CCA end + diphosphate

CTP:3'-cytidine-tRNA cytidylyltransferase activity
O:0052927 CTP:tRNA cytidylyltransferase activity
A tRNA precursor + 2 CTP <=> a tRNA with a 3' CC end + 2 diphosphate

This makes retrofitting annotations tricky. From the sound it, this could potentially be automated based on species.

[Harold J. Drabkin]

It appears the might have gotten split out
tRNA + CTP -> tRNA-C
Then
CTP + tRNA-C -> tRNA-CC
then
tRNA-CC + ATP -> tRNA-CCA
In some organisms one enzyme does steps 1-3, even though we know it is sequencial (ie, CCA is not made first and then added).

I just want to make sure that there is not a separate enzyme for rxn1 vs 2.(contacting my expert now), so that in some organsims it is 1+2 (again, sequentially).

[David Osumi-Sutherland]

Some cross-posting confusion going on here. Looking at paper more carefully.

[Harold J. Drabkin]

Yes, I just want to be sure that there is NOT a case that JUST the C is added.
It's just because I am wondering why this one was made to begin with
GO:0052927 CTP:tRNA cytidylyltransferase activity
Catalysis of the reaction: a tRNA precursor + CTP = a tRNA with a 3' cytidine end + diphosphate.

[David Osumi-Sutherland]

But if it's mechanistically the same reaction, then couldn't a single GO molecular function term could cover all the reactions?

That would certainly be easier for us.

Harold: Perhaps more efficient for you to take the ticket, given that you're getting some expert input?

Cheers,
David

[Harold J. Drabkin]

Peter: minimally two are needed. If we only had one to encompass all three, then there could be annotation error if someone were annotating the two enzymes that only add CC and only add the A. Only in eukaryotes would it be ok to say one enzyme did C1, C2, and then A1.