Inferred from Key Residues (IKR)

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  • IKR is a type of manually-curated evidence derived from sequence analysis, characterized by the lack of key sequence residues. All annotations that apply this evidence code should use the 'NOT' qualifier. This evidence code is used to annotate a gene product when, although homologous to a particular protein family, it has lost essential residues and is very unlikely to be able to carry out an associated function, participate in the expected associated process, or found in a certain location. This annotation statement can be supported by a published literature reference (e.g. a PubMed identifier) that has described the sequence analysis efforts, or by a GO Reference that describes the process a curator undertook to become sufficiently convinced of the sequence mutation. This evidence code is also referred to as IMR (inferred from Missing Residues).

Examples of IKR Usage

  • Curator-Determined IKR Annotation Example: Rat HPT (P06866) is homologous to serine proteases and contains a match to the peptidase S1 domain. However further sequence analysis by a curator looking at the the Peptidase S1B, active site established it has lost all essential catalytic residues, making it unable to carry out serine protease activity.
  • Curator-Determined IKR Annotation Example, Using PAINT :
  • Paper-Curated IKR Annotation Example: Ross,J., Jiang,H., Kanost,M.R. and Wang,Y. (2003) Serine proteases and their homologs in the Drosophila melanogaster genome: an initial analysis of sequence conservation and phylogenetic relationships. Gene 30;304:117-31 (PMID:12568721). The authors describe the determination of serine protease activity of proteins from the D. melanogaster S1 serine protease gene family, by determining the presence of conserved His, Asp, Ser catalytic triad residues in retrieved sequences. If all three residues were present in the conserved TAAHC, DIAL, and GDSGGP motifs, the sequence was considered to have serine protease activity. Any sequence lacking one of the key residues was identified as an a serine protease homolog, lacking proteolytic activity.
DB Object ID DB Object Symbol Qualifier GO ID DB:Reference Evidence Code With/From
UniProtKB:P06866 RatHPT NOT GO:0004252 (serine-type endopeptidase activity) GO_REF:0000047 IKR InterPro:IPR000126
FB:FBgn0033192 gene S1 NOT GO:0004252 (serine-type endopeptidase activity) PMID:12568721 IKR

Use of the With/From Field for IKR

  • Where an IKR annotation statement is made using a GO Reference, inclusion of an identifier in the With/From field that can indicate to the user the lacking residues (e.g. an alignment, domain or annotation rule identifier) is absolutely required.
  • In contrast, when an IKR annotation statement is supported by a published literature reference, a value in the With/From field is highly recommended, although not absolutely required.

When IKR Should NOT be Used

  • If, in a paper, an experiment is performed that demonstrates the lack of an activity, for example protein tyrosine phosphatase activity, then the curator should use the appropriate experimental evidence code and the NOT qualifier to annotate that gene/gene product.
  • If a paper supplies data that showed the active site was missing and additionally carried out an experimental assay to show lack of activity, it would be correct to create two annotation statements from this paper; both NOT IKR and NOT IDA.
  • CAUTION: Where curators make judgements of function using the IKR evidence code, they should be able to draw on some level of expertise regarding the protein family, as there will always be exceptions to the rule. For instance, Q9H4A3 (WNK1_HUMAN) is a good example where nature has confounded prediction; Cys250 is present instead of the conserved Lys which is expected to be an active site residue. However Lys233 appears to fulfill the required catalytic function.

Quality Control Checks

Evidence and Conclusion Ontology

ECO:0000320 phylogenetic determination of loss of key residues evidence used in manual assertion


Curator Guide to GO Evidence Codes

Gene Ontology website GO Evidence Codes list

Review Status

Last reviewed: April 6, 2018