1. An example of GO annotation from a primary paper GO Annotation Camp, July 2006 PMID:12972596 http://www.ebi.ac.uk/GOA

2. In this paper two proteins can be annotated to: UniProtKB accession: Q96T23UniProtKB accession: O60264 Loyola et al. provide experimental evidence for biological process (BP), molecular function (MF) and cellular component (CC) annotations for both Rsf-1 and hSNF2H. Rsf-1 and hSNF2H …from the gene/protein names found in the paper, you then need to find the corresponding sequence identifier suitable for your database …

3. NB: Rsf-1 and hSNF2H were previously shown to form the RSF complex, and have therefore already been annotated to the GO term: RSF complex ; GO:0031213 (in PMID: 9836642)

4. Starting on Page 6762, 2 nd paragraph in left column… Q96T23 ( Rsf-1) nucleus ; GO:0005634 IDA O60264 ( hSNF2H) nucleus ; GO:0005634 IDA NB: There are two sets of evidence that the subunits of RSF localize to the nucleus: 1/ the complex was purified from a nuclear fraction. 2/ antibody staining. Both of these pieces of information provide a direct assay for the subcellular localization of these proteins. For any given paper, if both the GO term and the evidence code are identical the annotation is only recorded once. To assign cellular component annotations….

5. Figure 2B: Subcellular localization of Rsf-1 and hSNF2H, visualised with specific antibodies. Both Rsf-1 and hSNF2H localize to the nucleus. Additionally, localization of hSNF2H is restricted to chromosomes during mitosis. NB: Using GO, it is not possible to state that hSNF2H is nuclear during interphase and restricted to a chromosome during metaphase. But both subcellular localizations can be attributed to hSNF2H with GO terms. (the temporal information is captured in UniProt separately)

6. Pg 6762, half-way down left hand column… … however when a GO term is made obsolete, the ‘comments’ often suggest alternative GO terms that may be appropriate to use. But the GO term ‘mitotic chromosome ; GO:0005708’ is obsolete.

7. hSNF2H: O60264 condensed chromosome ; GO:0000793 IDA ‘Open mitosis’ occurs in humans, so GO:0000793 can be used to annotate hSNF2H.

8. Can annotate with the GO molecular function ‘protein binding’ (GO:0005515) based on these assays… Pg 6762, half way down left hand column… To assign Molecular Function ‘protein binding’ annotations….

9. Figure 2C: Immunoprecipitation experiments support ‘protein binding’ GO annotations.

10. NB: When using the IPI code, include also a sequence identifier for the interacting protein in the ‘with’ field. For protein binding GO terms, always provide the reciprocal annotation. Rsf-1 ‘ with’ Q96T23 protein binding ; GO:0005515 IPI O60264 ALSO… hSNF2H O60264 protein binding ; GO:0005515 IPI Q96T23

11. Pg 6762, 2 nd paragraph in the right hand column … hSNF2H: O60264 ATPase activity ; GO:0016887 IDA To assign a Molecular Function ‘ATPase activity’ annotation…. Rsf-1: Q96T23 NOT ATPase activity ; GO:0016887 IDA

12. Use of the ‘NOT’ qualifier The ‘NOT’ qualifier is used when a GO term might otherwise be expected to apply to a gene product, but an experiment etc. proves otherwise. Because the recombinant RSF complex shows ATPase activity, it is informative to record for this paper (using the ‘NOT’ qualifier) that the Rsf-1 subunit of RSF does not possess this activity. For further documentation on the ‘NOT’ qualifier, refer to the GO web site at: http://www.geneontology.org/GO.annotation.shtml

13. Pg 6762, 2 nd paragraph in the right hand column … hSNF2H: O60264 ATPase activity ; GO:0016887 IDA To assign a Molecular Function ‘ATPase activity’ annotation…. Rsf-1: Q96T23 NOT ATPase activity ; GO:0016887 IDA

14. hSNF2H: ‘with’ O60264 ATP binding ; GO:0005524 IC GO:0016887 hSNF2H: O60264 ATPase activity ; GO:0016887 IDA

15. IC: Inferred by curator Such an annotation is created where there is no direct experimental evidence or author statement in the paper, but where an annotation can be reasonably inferred by a curator from other GO annotations (for which evidence is available). In this paper the authors show that hSNF2H has ATPase activity. It can therefore be reasonably inferred that hSNF2H can bind ATP. The IC evidence code is used to support the ‘ATP binding’ annotation to show that a curator has made this inference. For evidence code documentation, see: http://www.geneontology.org/GO.evidence.shtml

16. To assign a Biological Process GO annotation…. Page 6762, half way down right hand column… hSNF2H: O60264 chromatin assembly ; GO:0031497 IDA Rsf-1: Q96T23 chromatin assembly ; GO:0031497 IDA

17. Rsf-1: Q96T23 histone binding ; GO:0042393 NAS hSNF2H: O60264 contributes_to histone binding ; GO:0042393 NAS Page 6762, 3 rd paragraph of right hand column… To assign a Molecular Function ‘histone binding’ annotation…. NB: When data is not shown in a paper but the results are still described, the NAS evidence code is generally used to support GO annotation (subject to judgement by the annotator). If time allows, we would also annotate Ref (13) to see if we can assign histone binding annotation to Rsf-1 with an experimental evidence code.

18. Use of the ‘contributes_to’ qualifier A gene product that is part of a complex can be annotated to the molecular function terms that describe the action of the whole complex (i.e. annotating 'to the potential of the complex‘) It is a way to capture information about what a complex does in the absence of database objects and identifiers representing complexes. Molecular function annotations to subunits that do not possess the activity of the complex themselves must include the qualifier ‘contributes_to’. All gene products annotated using 'contributes_to' must also be annotated to a cellular component term representing the complex that possesses the activity. hSNF2H already has the cellular component annotation RSF complex ; GO:0031213, from annotation to another paper and therefore a ‘contributes_to’ histone binding annotation can be made. For further documentation on the ‘contributes_to’ qualifier, refer to the GO web site at: http://www.geneontology.org/GO.annotation.shtml

19. hSNF2H: O60264 DNA binding ; GO:0003677 IDA Page 6762, last paragraph of right hand column… Page 6764, bottom of the page, left hand column… To assign a Molecular Function ‘DNA binding’ annotation….

20. The DNA binding properties of Rsf-1 are slightly more complicated…… Page 6765, left hand column…

21. Rsf-1 Q96T23 negative regulation of DNA binding ; GO:0043392 IDA At the time of curation a term for ‘negative regulation of DNA binding’ did not exist. Therefore a SourceForge item was made to request that this new term be added to GO.

22. Rsf-1‘with’ CC nucleus ; GO:0005634IDA MF protein binding ; GO:0005515IPI O60264 histone binding ; GO:0042393NAS NOT ATPase activity ; GO:0016887 IDA BP chromatin assembly ; GO:0031497 IDA negative regulation of DNA binding ; GO:0043392 IDA hSNF2H CC nucleus ; GO:0005634 IDA condensed chromosome ; GO:0000793 IDA MF protein binding ; GO:0005515 IPI Q96T23 ATPase activity ; GO:0016887 IDA ATP binding ; GO:0005524 IC GO:0016887 DNA binding ; GO:0003677 IDA contributes_to histone binding ; GO:0042393 NAS * BP chromatin assembly ; GO:0031497 IDA Summary of GO annotation for PMID:12972596 * RSF complex ; GO:0031213 annotated from PMID:9836642

23. Q96T23 (Rsf-1)

24. O60264 (hSNF2H)