Curator Guide: Enzymes and Reactions

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This guide is for editors adding molecular function terms to represent enzyme reactions.

There are five websites that are particularly useful when adding reaction terms. These are:

For chemical names, one should consult ChEBI. RHEA is particularly useful because it gives EC reactions using ChEBI chemicals.


General Rules and Things of Note

Enzyme Commission

The Enzyme Commission names and categorises enzymes, i.e. physical entities, whereas GO is interested in the various reactions that the enzyme performs. In the same way that a gene product may participate in a number of different processes, it may catalyse a number of different reactions; the ontology should contain each reaction, and the job of the annotator is to mark which reactions a certain gene product catalyses. A single enzyme may perform a number of different reactions, and it is also possible for several different EC enzymes to perform the same reaction.

This means that there is not a 1:1 correspondence between EC numbers and GO reaction terms.

There are a number of websites that mirror the EC data; my favourite is IntEnz as it shows the reactions from RHEA, so you are saved the trouble of having to find out what the ChEBI names for the reaction participants are.


MetaCyc

At present, MetaCyc reactions are associated with one EC number, so if two different EC enzymes catalyse the same reaction, there will be two MetaCyc reactions, one for each EC number.


KEGG

KEGG makes reactions independent of the EC number; you can look up an EC number and see the reactions that the enzyme performs (e.g. EC:1.1.1.21), or you can look up a reaction and see which EC enzymes perform that reaction (e.g. R01036). Nifty!


Reactome

Reactome currently provide mappings of their terms to GO terms, so they do the work for us!


Precise vs. Imprecise EC Numbers

GO has terms that represent the categories used by EC. These have EC xrefs of the form EC:n, EC:n.n and EC:n.n.n (where n is a number).

For reactions where the enzyme has not yet been added to EC, but it can be put into one of the EC categories, the xref should be of the form EC:n.n.n.-, i.e. ending with a dash.


NAD(P)

Here is the (September 2013) view from the IUBMB committee (Keith Tipton and other members clarified this) after canvasing by Kristian Axelson and Alan Bridge:

Our 'classification Rules (on both websites) clearly state in rule 18: "Where the enzyme can use either coenzyme, this should be indicated by writing NAD(P)+".

For further info see rule 18 on systematic names at http://www.chem.qmul.ac.uk/iubmb/enzyme/rules.html

So the meaning is really: "the enzyme can use both", rather than "the reaction may contain either".


 alditol + NAD(P)+ = aldose + NAD(P)H + H+

means that the enzyme performs

 alditol + NAD+ = aldose + NADH + H+

AND

 alditol + NADP+ = aldose + NADPH + H+

One EC number, multiple reactions

There are a number of cases where an enzyme can catalyse a set of reactions. These may or may not be specified by EC, but KEGG and MetaCyc will often show additional reactions. Similarly, there are often different EC enzymes that will catalyse the same reaction. A good example of this overlap is found in EC:1.5.3.13, 14, 15, 16, and 17. Looking at IntEnz, there are four reactions for EC:1.5.3.17; if we then look at EC:1.5.3.16, we can see that one of the reactions from EC:1.5.3.17 can be catalysed by this enzyme, too. KEGG shows this data more clearly; viewing all the reactions for EC:1.5.3.17 (click 'Show all' on the enzyme data page), each reaction has the EC numbers of enzymes that can catalyse it listed. MetaCyc also lists a number of reactions for each EC number.

Example 1: epi-cedrol synthase

Add a term for EC 4.2.3.39, epi-cedrol synthase

  • Check the reaction does not exist in GO by searching on the name, EC number and the reactants. I searched for 'epicedrol' and 'epi-cedrol'.
  • Look up the reaction in EC (using IntEnz), MetaCyc and KEGG.
2-trans,6-trans-farnesyl diphosphate + H2O <=> epi-cedrol + diphosphate
(2E,6E)-farnesyl diphosphate + H2O <=> 8-epi-cedrol + diphosphate
trans,trans-Farnesyl diphosphate + H2O <=> 8-epi-Cedrol + Diphosphate

Check against the RHEA reaction, RHEA:26118 (linked from IntEnz) so that we can be sure we're using the correct nomenclature.

Names and synonyms: KEGG and EC both give us "(2E,6E)-farnesyl-diphosphate diphosphate-lyase (8-epi-cedrol-forming)", which is the systematic name, according to EC. We also have "8-epicedrol synthase" and "epicedrol synthase".

Parentage: find the GO term for the category EC:4.2.3; if any of the children are relevant, use them as the parent.

name: epi-cedrol synthase activity
def: "Catalysis of the reaction: 2-trans,6-trans-farnesyl diphosphate + H2O = epi-cedrol + diphosphate." [RHEA:26118]
synonym: "(2E,6E)-farnesyl-diphosphate diphosphate-lyase (8-epi-cedrol-forming) activity" EXACT systematic_synonym [EC:4.2.3.39]
synonym: "8-epicedrol synthase activity" EXACT []
synonym: "epicedrol synthase activity" EXACT []
xref: EC:4.2.3.39
xref: MetaCyc:RXN-10004
xref: KEGG:R09140
xref: RHEA:26118
is_a: GO:0016838 ! carbon-oxygen lyase activity, acting on phosphates


Example 2: farnesol kinase

From SourceForge:

 definition: farnesol + an NTP = farnesol phosphate + an NDP
 EC: 2.7.1.-
 One example of a more specific case of this is: MetaCyc RXN-11625
 
 PMID 21395888
 PMID 10557276
 
 NARROW synonym: trans,trans-farnesol kinase
 NARROW synonym: 2-trans, 6-trans-farnesol kinase
  • Look up the MetaCyc reaction. It's
2-trans,-6-trans-farnesol + CTP = 2-trans,-6-trans-farnesyl monophosphate + CDP + H+
  • Search GO, EC, KEGG and RHEA for farnesol. No results for reactions of a similar form.
  • Checking the literature references, it is not clear whether the farnesol reactions are limited to the 2-trans,6-trans isomer, so we'll refer to 'farnesol' in the reaction.
  • ChEBI searches for farnesol phosphates turn up a blank; however, "farnesyl phosphate" is a parent term for "farnesyl diphosphate" so we should use the name "farnesyl monophosphate" instead of "farnesol phosphate" to refer to the reaction product.
  • Parentage: MetaCyc gives an EC ref of 2.7.1.- for RXN-11625; this corresponds to GO:0016773. We can have a look at the ChEBI hierarchy for "farnesyl phosphate" to get some hints as to whether there may be any generic terms under GO:0016773, but there don't seem to be any. (N.b. a 'prenol kinase' term was later added which would be a more appropriate parent)
  • Reaction equation: NTP and NDP are referred to in ChEBI as nucleoside triphosphate and nucleoside diphosphate.
name: farnesol kinase activity
def: "Catalysis of the reaction: farnesol + nucleoside triphosphate = farnesyl monophosphate + nucleoside diphosphate." [MetaCyc:RXN-11625]
synonym: "trans,trans-farnesol kinase activity" NARROW
xrefs: EC:2.7.1.-
is_a: GO:0016773 ! phosphotransferase activity, alcohol group as acceptor


  • Add the MetaCyc reaction cited as a child of this new term. I gave it the name "2-trans,-6-trans-farnesol kinase activity" to reflect the specific substrate.


Example 3: phosphomethylethanolamine N-methyltransferase activity

From SourceForge:

 Def: Catalysis of the reaction: phosphomethylethanolamine (PMEA) + AdoMet -> phosphodimethylethanolamine
 Ref: GOC:tb
 PMID 20650897


Searching for the enzyme name brings up no results in GO, EC, MetaCyc and KEGG, so let's look up the reaction instead.

Look up all three compounds mentioned in MetaCyc and KEGG.

Check the reactions for these compounds.

  • KEGG: R06868 looks like a match:
 S-Adenosyl-L-methionine + N-Methylethanolamine phosphate <=>
 S-Adenosyl-L-homocysteine + Phosphodimethylethanolamine
 N-methylethanolamine phosphate + S-adenosyl-L-methionine <=>
 N-dimethylethanolamine phosphate + S-adenosyl-L-homocysteine + H+
  • Check that N-dimethylethanolamine phosphate (from the MetaCyc reaction) is also known as phosphodimethylethanolamine
    • phosphodimethylethanolamine is a synonym on the MetaCyc compound page; the KEGG compound ID C13482 matches that in the KEGG reaction
    • If in doubt, search for the compound in ChEBI and check the synonyms.
  • MetaCyc states that the reaction is one of three catalysed by EC:2.1.1.103, so go to IntEnz and look up 2.1.1.103. Although the comments mention subsequent reactions, the reaction list doesn't, so we will use the more generic EC:2.1.1.- as a reference.
  • Get the ChEBI names for the substances and generate a balanced equation. Check to see if the reaction is in Rhea. I looked at the automatic xrefs for N-methylethanolamine phosphate in ChEBI and clicked on the Rhea xrefs. RHEA:25322 is a match! Checking the xrefs for the Rhea reaction, they match the reactions in KEGG and MetaCyc that we found earlier.
  • Term name: a quick Google search reveals that 'phosphomethylethanolamine N-methyltransferase' appears to be the most common name for this term.
  • Synonyms: added the KEGG name for the reaction as an exact synonym with the scope set as 'systematic synonym'; also added a synonym using the ChEBI name for the chemical instead of phosphomethylethanolamine.
  • Term parentage: this term can go under N-methyltransferase activity.
name: phosphomethylethanolamine N-methyltransferase activity
def: "Catalysis of the reaction: N-methylethanolamine phosphate + S-adenosyl-L-methionine = N,N-dimethylethanolamine phosphate + S-adenosyl-L-homocysteine + H(+)." [RHEA:25322, KEGG:R06868, MetaCyc:RXN-5642]
synonym: "N-methylethanolamine phosphate N-methyltransferase activity" EXACT
synonym: "S-adenosyl-L-methionine:methylethanolamine phosphate N-methyltransferase activity" EXACT systematic_synonym [KEGG:R06868]
xref: EC:2.1.1.-
xref: KEGG:R06868
xref: MetaCyc:RXN-5642
xref: RHEA:25322
is_a: GO:0008170 ! N-methyltransferase activity


Example 4: updating EC:1.5.3.11, a transferred EC entry

From EC:

Transferred entry: polyamine oxidase. Now included with EC 1.5.3.13 N1-acetylpolyamine oxidase,
EC 1.5.3.14 polyamine oxidase (propane-1,3-diamine-forming), EC 1.5.3.15 N8-acetylspermidine
oxidase (propane-1,3-diamine-forming), EC 1.5.3.16 spermine oxidase and EC 1.5.3.17 non-specific
polyamine oxidase

This is a tricky entry as there is a lot of overlap between the reactions that each enzyme catalyses. The best way to handle it is to copy out all the reactions (either from IntEnz or KEGG) and then see which are duplicated. E.g.

 EC:1.5.3.13:
[RHEA:25815] N1-acetylspermidine + H2O + O2 <=> 3-acetamidopropanal + H2O2 + putrescine 
[RHEA:25803] N1-acetylspermine + H2O + O2 <=> 3-acetamidopropanal + H2O2 + spermidine 
[RHEA:25871] N1,N12-diacetylspermine + H2O + O2 <=> 3-acetamidopropanal + N1-acetylspermidine + H2O2 
[RHEA:25811] H2O + O2 + spermidine <=> 3-aminopropanal + H2O2 + putrescine 
[RHEA:25807] H2O + O2 + spermine <=> 3-aminopropanal + H2O2 + spermidine 
EC:1.5.3.16:
[RHEA:25807] H2O + O2 + spermine <=> 3-aminopropanal + H2O2 + spermidine 
EC:1.5.3.17 
[RHEA:25803] N1-acetylspermine + H2O + O2 <=> 3-acetamidopropanal + H2O2 + spermidine 
[RHEA:25807] H2O + O2 + spermine <=> 3-aminopropanal + H2O2 + spermidine 
[RHEA:25811] H2O + O2 + spermidine <=> 3-aminopropanal + H2O2 + putrescine 
[RHEA:25815] N1-acetylspermidine + H2O + O2 <=> 3-acetamidopropanal + H2O2 + putrescine 

From these lists, we can see that RHEA:25807 will have EC refs 1.5.3.13, 1.5.3.16 and 1.5.3.17; RHEA:25815 will have EC refs 1.5.3.13 and 1.5.3.17; and so on. The KEGG reaction display makes it easier to check which reactions are linked with which EC numbers once you have figured out the correspondence between RHEA IDs and KEGG IDs. KEGG also provides names for the reactions; there was one case where a reaction name clashed with an existing GO MF term, so I made the new term name more specific whilst keeping to the nomenclature conventions used by the other terms.

There ended up being a lot of new terms created; here's a sample:

 name: spermine:oxygen oxidoreductase (spermidine-forming) activity
 def: "Catalysis of the reaction: H(2)O + O(2) + spermine = 3-aminopropanal + H(2)O(2) + spermidine." [RHEA:25807]
 xref: EC:1.5.3.13
 xref: EC:1.5.3.16
 xref: EC:1.5.3.17
 xref: KEGG:R09076
 xref: MetaCyc:1.5.3.17-RXN
 xref: MetaCyc:RXN-9015
 xref: RHEA:25807 "H(2)O + O(2) + spermine = 3-aminopropanal + H(2)O(2) + spermidine"
 
 name: spermidine:oxygen oxidoreductase (3-aminopropanal-forming) activity
 def: "Catalysis of the reaction: H(2)O + O(2) + spermidine = 3-aminopropanal + H(2)O(2) + putrescine." [RHEA:25811]
 xref: EC:1.5.3.13
 xref: EC:1.5.3.17
 xref: KEGG:R09077
 xref: MetaCyc:RXN-10461
 xref: MetaCyc:RXN-12089
 xref: RHEA:25811 "H(2)O + O(2) + spermidine = 3-aminopropanal + H(2)O(2) + putrescine"
 
 name: N1-acetylspermine:oxygen oxidoreductase (3-acetamidopropanal-forming) activity
 def: "Catalysis of the reaction: H(2)O + N(1)-acetylspermine + O(2) = 3-acetamidopropanal + H(2)O(2) + spermidine." [RHEA:25803]
 xref: EC:1.5.3.13
 xref: EC:1.5.3.17
 xref: KEGG:R03899
 xref: MetaCyc:RXN-12090
 xref: MetaCyc:RXN-9940
 xref: RHEA:25803 "H(2)O + N(1)-acetylspermine + O(2) = 3-acetamidopropanal + H(2)O(2) + spermidine"
 
 name: N1-acetylspermidine:oxygen oxidoreductase (3-acetamidopropanal-forming) activity
 def: "Catalysis of the reaction: H(2)O + N(1)-acetylspermidine + O(2) = 3-acetamidopropanal + H(2)O(2) + putrescine." [RHEA:25815]
 xref: EC:1.5.3.13
 xref: EC:1.5.3.17
 xref: KEGG:R09074
 xref: MetaCyc:RXN-12091
 xref: MetaCyc:RXN-9942
 xref: RHEA:25815

There were also extra reactions in KEGG and MetaCyc that weren't in the EC listings; whether you add these or not depends on whether the person requesting the terms has asked for them and/or whether you want to add them.


Multi-Step Enzyme Reactions

  • When an enzyme reaction is multi-step, but with just one EC number, add one GO term for the overall reaction, and add a comment to say the reaction is multi-step (and whether one of the steps is spontaneous).
  • Comment: This is a multi-step reaction, with a spontaneous second step.

E.g. EC:1.14.13.109: http://www.chem.qmul.ac.uk/iubmb/enzyme/EC1/14/13/109.html

Accepted name: abieta-7,13-dien-18-ol hydroxylase
Reaction: abieta-7,13-dien-18-ol + NADPH + H+ + O2 = abieta-7,13-dien-18-al + NADP+ + 2 H2O (overall reaction)
(1a) abieta-7,13-dien-18-ol + NADPH + H+ + O2 = abieta-7,13-dien-18,18-diol + + NADP+ + H2O
(1b) abieta-7,13-dien-18,18-diol = abieta-7,13-dien-18-al + H2O (spontaneous)
  • An update on this issue (from Becky, 16.10.2012):

We decided that if the enzyme catalyzed all steps, and if all steps always occurred together, then we'd add GO terms for the separate steps as HAS_PART children of the overall reaction. The problem came with naming the individual steps... the plan was to take the names direct from KEGG, but KEGG don't always name the separate steps. I emailed them about it but haven't heard back from them yet:

Some of the background is at the top of this SF item: https://sourceforge.net/tracker/?func=detail&aid=3510070&group_id=36855&atid=440764

So at the moment, most GO terms have been added with the definition specifying that it's a 2/3/multi-step reaction (E.g. GO:0036209), ready for HAS_PART children to be added when we've sorted out how to name them.