celM

id: P55742
gene_symbol: celM
aliases:
  - CelM
  - TET aminopeptidase
  - M42 aminopeptidase
product_type: PROTEIN
status: DRAFT
taxon:
  id: NCBITaxon:203119
  label: Acetivibrio thermocellus
description: CelM is a putative aminopeptidase belonging to the M42 peptidase family,
  originally misannotated as an endoglucanase (hence the "cel" nomenclature). Despite
  its misleading name, CelM has been experimentally characterized as a metalloaminopeptidase
  with leucine aminopeptidase (LAP) activity, requiring cobalt ions (Co2+) as a cofactor.
  The protein forms an active dodecameric complex and preferentially cleaves nonpolar
  aliphatic L-amino acid residues from the N-terminus of peptide substrates, with
  L-leucine-pNA being the optimal substrate. CelM shows no significant cellulase
  activity despite earlier erroneous reports. The protein does not contain a dockerin
  domain and is therefore not a component of the cellulosome complex. Its biological
  role may involve protein turnover or peptide processing in the cytoplasm of A.
  thermocellus.
existing_annotations:
- term:
    id: GO:0004177
    label: aminopeptidase activity
  evidence_type: IEA
  original_reference_id: GO_REF:0000043
  review:
    summary: This annotation is correct and supported by experimental evidence from
      Dutoit et al. (2012) PMID:23226342, who demonstrated that CelM has leucine
      aminopeptidase (LAP) activity using L-leucine-pNA as substrate. The protein
      preferentially hydrolyzes nonpolar aliphatic L-amino acid-pNA substrates at
      the N-terminus.
    action: ACCEPT
    reason: CelM has been experimentally demonstrated to function as an aminopeptidase.
      Although the annotation is derived from keyword mapping (IEA), it accurately
      reflects the experimentally determined function of the protein. This is a core
      molecular function of the enzyme.
    additional_reference_ids:
    - PMID:23226342
    supported_by:
    - reference_id: UniProt:P55742
      supporting_text: Belongs to the peptidase M42 family.
- term:
    id: GO:0006508
    label: proteolysis
  evidence_type: IEA
  original_reference_id: GO_REF:0000043
  review:
    summary: This annotation is correct as CelM participates in proteolytic degradation
      through its aminopeptidase activity. The protein hydrolyzes peptide bonds at
      the N-terminus of peptides.
    action: ACCEPT
    reason: Aminopeptidases are proteolytic enzymes that participate in proteolysis
      by cleaving N-terminal amino acid residues from peptides. This is the appropriate
      biological process term for the enzymatic activity of CelM.
    additional_reference_ids:
    - PMID:23226342
    supported_by:
    - reference_id: UniProt:P55742
      supporting_text: Belongs to the peptidase M42 family.
- term:
    id: GO:0008233
    label: peptidase activity
  evidence_type: IEA
  original_reference_id: GO_REF:0000043
  review:
    summary: This annotation is correct but represents a parent term of the more
      specific aminopeptidase activity annotation. CelM is a peptidase of the M42
      metallopeptidase family.
    action: ACCEPT
    reason: This is a valid annotation as CelM is indeed a peptidase. However, the
      more specific term GO:0004177 (aminopeptidase activity) or GO:0070006 (metalloaminopeptidase
      activity) is more informative. This general term is acceptable as an IEA annotation
      alongside the more specific annotations.
    additional_reference_ids:
    - PMID:23226342
    supported_by:
    - reference_id: UniProt:P55742
      supporting_text: Belongs to the peptidase M42 family.
- term:
    id: GO:0008237
    label: metallopeptidase activity
  evidence_type: IEA
  original_reference_id: GO_REF:0000043
  review:
    summary: This annotation is correct. CelM is an M42 family metallopeptidase that
      requires divalent metal cations (specifically Co2+ for optimal activity) for
      catalysis. The metal ions are essential for the enzymatic mechanism.
    action: ACCEPT
    reason: CelM is a metallopeptidase that requires metal ions for activity. EDTA
      chelation inhibits activity, confirming the metal-dependence of the enzyme.
      The annotation correctly reflects the catalytic mechanism of the enzyme.
    additional_reference_ids:
    - PMID:23226342
    supported_by:
    - reference_id: UniProt:P55742
      supporting_text: Binds 2 divalent metal cations per subunit.
- term:
    id: GO:0016787
    label: hydrolase activity
  evidence_type: IEA
  original_reference_id: GO_REF:0000043
  review:
    summary: This annotation is correct but very general. CelM is a hydrolase that
      catalyzes the hydrolysis of peptide bonds. This is a parent term of the more
      specific peptidase and aminopeptidase activity terms.
    action: ACCEPT
    reason: While correct, this is the most general term in the hierarchy. The more
      specific annotations (aminopeptidase activity, metallopeptidase activity) are
      more informative. This annotation is acceptable as part of the IEA annotation
      set derived from keyword mapping, but provides limited functional insight beyond
      the more specific terms.
    supported_by:
    - reference_id: UniProt:P55742
      supporting_text: Belongs to the peptidase M42 family.
- term:
    id: GO:0046872
    label: metal ion binding
  evidence_type: IEA
  original_reference_id: GO_REF:0000043
  review:
    summary: This annotation is correct. CelM binds divalent metal cations which
      are essential for its catalytic activity. UniProt reports binding of 2 divalent
      metal cations per subunit at defined binding sites.
    action: ACCEPT
    reason: CelM requires divalent metal cations for activity. UniProt documents
      metal binding sites at residues 65, 168, 199, 221, and 307. While cobalt provides
      optimal activity in vitro, the in vivo metal cofactor may differ. The general
      metal ion binding term is appropriate given this uncertainty.
    additional_reference_ids:
    - PMID:23226342
    supported_by:
    - reference_id: UniProt:P55742
      supporting_text: Binds 2 divalent metal cations per subunit.
- term:
    id: GO:0070006
    label: metalloaminopeptidase activity
  evidence_type: IDA
  original_reference_id: PMID:23226342
  review:
    summary: This is the most appropriate and specific molecular function term for
      CelM. The protein is an M42 family metalloaminopeptidase that cleaves N-terminal
      amino acids using a metal-dependent catalytic mechanism.
    action: NEW
    reason: This term should be added as a new annotation based on the experimental
      characterization by Dutoit et al. (2012). CelM is a metalloaminopeptidase that
      requires metal ions for catalysis and cleaves N-terminal amino acids from peptide
      substrates. This is more specific than both aminopeptidase activity and metallopeptidase
      activity.
    additional_reference_ids:
    - PMID:23226342
    supported_by:
    - reference_id: UniProt:P55742
      supporting_text: Belongs to the peptidase M42 family.
references:
- id: GO_REF:0000043
  title: Gene Ontology annotation based on UniProtKB/Swiss-Prot keyword mapping
  full_text_unavailable: false
  findings:
  - statement: Keywords for Aminopeptidase, Hydrolase, Metal-binding, Metalloprotease,
      and Protease were correctly used to infer GO annotations for molecular function.
- id: PMID:23226342
  title: Functional characterization of two M42 aminopeptidases erroneously annotated
    as cellulases.
  findings:
  - statement: CelM from Clostridium thermocellum was experimentally characterized
      as an M42 family aminopeptidase, not a cellulase as previously annotated.
    supporting_text: "some members have been annotated as cellulases because of their homology with CelM, formerly described as an endoglucanase of Clostridium thermocellum"
  - statement: The protein shows no significant cellulase activity on CMC and cellobiose.
    supporting_text: "No significant endoglucanase activity was measured, either for TmPep1050 or CelM"
  - statement: CelM has robust leucine aminopeptidase activity with L-leucine-pNA
      as the optimal substrate.
    supporting_text: "Both enzymes were shown to catalyze hydrolysis of nonpolar aliphatic L-amino acid-pNA substrates, the L-leucine derivative appearing as the best substrate"
  - statement: Cobalt ions (Co2+) are required for maximal activity.
    supporting_text: "Addition of cobalt ions enhanced the activity of both enzymes significantly"
  - statement: EDTA inhibits activity, confirming metal-dependence.
    supporting_text: "both the chelating agent EDTA and bestatin, a specific inhibitor of metalloaminopeptidases, proved inhibitory"
  - statement: Bestatin inhibits CelM, consistent with metalloaminopeptidase function.
    supporting_text: "bestatin, a specific inhibitor of metalloaminopeptidases, proved inhibitory"
- id: UniProt:P55742
  title: UniProt entry for CelM/Putative aminopeptidase
  full_text_unavailable: false
  findings:
  - statement: Classified as Peptidase M42 family member.
    supporting_text: Belongs to the peptidase M42 family.
    reference_section_type: DATABASE_ENTRY
  - statement: Binds 2 divalent metal cations per subunit.
    supporting_text: Binds 2 divalent metal cations per subunit.
    reference_section_type: DATABASE_ENTRY
core_functions:
- description: Metalloaminopeptidase that cleaves N-terminal amino acids from peptides
    using a metal-dependent catalytic mechanism, with preference for nonpolar aliphatic
    amino acids like leucine. Requires cobalt ions for optimal activity in vitro
    and forms active dodecameric complexes. Not a cellulase despite the gene name.
  molecular_function:
    id: GO:0070006
    label: metalloaminopeptidase activity
  directly_involved_in:
  - id: GO:0006508
    label: proteolysis
  supported_by:
  - reference_id: UniProt:P55742
    supporting_text: Belongs to the peptidase M42 family.
suggested_questions:
- question: What is the physiological metal cofactor of CelM in vivo? While cobalt
    provides optimal activity in vitro, the in vivo cofactor may be different.
- question: What is the biological role of CelM in A. thermocellus? Is it involved
    in protein turnover, processing of cellulosomal components, or another cellular
    function?
- question: Does CelM have any role related to the cellulosome, or is it entirely
    a cytoplasmic housekeeping enzyme?
suggested_experiments:
- description: Determine the native metal cofactor of CelM through metal content
    analysis of purified enzyme from A. thermocellus.
- description: Construct celM deletion mutant to assess phenotypic effects and identify
    physiological role.
- description: Identify physiological substrates through proteomics or peptide library
    screening.
- description: Determine if CelM is cytoplasmic or secreted, and if it has any association
    with cellulosome components.