IKR: Inferred from Key Residues

Updated May 2, 2012

  • 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. Where an IKR annotation statement is made using a GO Reference, inclusion of an identifier in the 'with/from' column of the annotation format 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 required. This evidence code is also referred to as IMR (inferred from Missing Residues).

Examples where the IKR evidence code should be used:

  • 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 : Curators determined that Drosophila neuroligin protein does not have carboxylesterase activity, based on phylogeny-based evidence. The Panther identifier in the 'with/from' field links out to an evidence record citing annotation data from orthologous gene products, supporting the annotation statement.
  • 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








Evidence Code



... P06866 RatHPT NOT GO:0004252serine-type endopeptidase activity GO_REF:0000047 IKR InterPro:IPR000126 ...
... P06866 neuroligin NOT GO:0004091carboxylesterase activity GO_REF:0000033 IKR PANTHER:PTHR11559_AN146 ...
... FB:FBgn0033192 gene S1 NOT GO:0004252serine-type endopeptidase activity PMID:12568721 IKR   ...

Examples where the IKR evidence code should not be used:

  • If there is experimental evidence available from a publication to support a NOT-evidenced annotation. In such instances, the curator should make the IDA, IMP or EXP NOT-qualified annotation based on the experimental evidence. 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 functionning 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; Cys-250 is present instead of the conserved Lys which is expected to be an active site residue. However Lys-233 appears to fulfill the required catalytic function.