Existing Annotations Review

GO Term	Evidence	Action	Reason
GO:0008734 L-aspartate oxidase activity	IEA GO_REF:0000120	ACCEPT	Summary: This is the specific catalytic activity supported for AO_0. Reason: UniProt and family assignment consistently identify AO_0 as an L-aspartate oxidase rather than a generic oxidoreductase. Supporting Evidence: file:NICAT/NaAO2_candidate_AO_0/NaAO2_candidate_AO_0-notes.md UniProt curates A0A1J6KBX2 as an L-aspartate oxidase/NadB-family flavoprotein that catalyzes oxidation of L-aspartate to iminoaspartate, linking this candidate to the upstream NAD branch that feeds pyridine alkaloid biosynthesis.
GO:0009435 NAD+ biosynthetic process	IEA GO_REF:0000120	ACCEPT	Summary: AO_0 belongs in NAD biosynthesis. Reason: Aspartate oxidase catalyzes the upstream reaction that feeds de novo NAD biosynthesis, which is the ancestral source pathway for the pyridine branch. Supporting Evidence: file:NICAT/NaAO2_candidate_AO_0/NaAO2_candidate_AO_0-notes.md UniProt curates A0A1J6KBX2 as an L-aspartate oxidase/NadB-family flavoprotein that catalyzes oxidation of L-aspartate to iminoaspartate, linking this candidate to the upstream NAD branch that feeds pyridine alkaloid biosynthesis.
GO:0009507 chloroplast	IEA GO_REF:0000044	KEEP AS NON CORE	Summary: Chloroplast localization is plausible and should be retained as non-core context. Reason: The location is supported in UniProt, but the main curation question for this candidate is paralog assignment within the pathway rather than subcellular detail. Supporting Evidence: file:NICAT/NaAO2_candidate_AO_0/NaAO2_candidate_AO_0-uniprot.txt CC -!- SUBCELLULAR LOCATION: Plastid, chloroplast
GO:0016491 oxidoreductase activity	IEA GO_REF:0000002	MARK AS OVER ANNOTATED	Summary: This parent oxidoreductase term is true but not specific enough. Reason: The more informative catalytic term GO:0008734 already captures the actual chemistry of AO_0. Supporting Evidence: file:NICAT/NaAO2_candidate_AO_0/NaAO2_candidate_AO_0-notes.md UniProt curates A0A1J6KBX2 as an L-aspartate oxidase/NadB-family flavoprotein that catalyzes oxidation of L-aspartate to iminoaspartate, linking this candidate to the upstream NAD branch that feeds pyridine alkaloid biosynthesis.

Core Functions

AO_0 is an L-aspartate oxidase candidate in the duplicated NAD-derived branch that supplies pyridine precursor chemistry to nicotine biosynthesis.

Molecular Function:

L-aspartate oxidase activity

Directly Involved In:

NAD+ biosynthetic process

Supporting Evidence:

file:NICAT/NaAO2_candidate_AO_0/NaAO2_candidate_AO_0-notes.md
AO_0 is therefore a plausible NaAO2 candidate, but the current public record does not distinguish it cleanly from AO_1 as the nicotine-pathway-specialized copy.

References

GO_REF:0000002

Gene Ontology annotation through association of InterPro records with GO terms

GO_REF:0000044

Gene Ontology annotation based on UniProtKB/Swiss-Prot Subcellular Location vocabulary mapping, accompanied by conservative changes to GO terms applied by UniProt

GO_REF:0000120

Combined Automated Annotation using Multiple IEA Methods

file:NICAT/NaAO2_candidate_AO_0/NaAO2_candidate_AO_0-uniprot.txt

UniProt entry A0A1J6KBX2 for Nicotiana attenuata AO_0

AO_0 is annotated as an L-aspartate oxidase in NAD precursor metabolism

"DE RecName: Full=L-aspartate oxidase"
UniProt places AO_0 in chloroplast-localized NAD biosynthetic chemistry

"CC -!- SUBCELLULAR LOCATION: Plastid, chloroplast"

file:NICAT/NaAO2_candidate_AO_0/NaAO2_candidate_AO_0-notes.md

NaAO2 AO_0 candidate notes

AO_0 is a plausible upstream pyridine-branch nicotine candidate

"UniProt curates A0A1J6KBX2 as an L-aspartate oxidase/NadB-family flavoprotein that catalyzes oxidation of L-aspartate to iminoaspartate, linking this candidate to the upstream NAD branch that feeds pyridine alkaloid biosynthesis."
AO_0 versus AO_1 remains an explicit paralog-resolution problem

"AO_0 is therefore a plausible NaAO2 candidate, but the current public record does not distinguish it cleanly from AO_1 as the nicotine-pathway-specialized copy."

Suggested Experiments

Experiment: Compare AO_0 and AO_1 transcript abundance across roots, leaves, and induction time courses relevant to nicotine biosynthesis.

Hypothesis: One AO paralog is preferentially associated with nicotine-producing root tissue.

Type: expression profiling

Experiment: Create single-paralog knockouts and measure NAD intermediates, nicotinic acid branch metabolites, and nicotine accumulation.

Hypothesis: The pathway-specialized AO paralog will have a stronger effect on nicotine precursor supply.

Type: genetic perturbation plus metabolite profiling

Deep Research

OpenAI

(NaAO2_candidate_AO_0-deep-research-openai.md)

NaAO2_candidate_AO_0-deep-research-openai OpenAI gpt-4.1 5 citations 2026-04-05T13:06:47.909171

The L-aspartate oxidase encoded by the NaAO2_candidate_AO_0 gene in Nicotiana attenuata plays a fundamental role in the de novo biosynthesis of NAD⁺, initiating the pathway by oxidizing L-aspartate to iminoaspartate. While specific studies on this enzyme in Nicotiana attenuata are limited, insights from model organisms like Arabidopsis thaliana provide a framework for understanding its function, regulation, and structural characteristics. Further research is needed to elucidate the specific roles and regulatory mechanisms of LASPO in Nicotiana attenuata, particularly concerning its involvement in secondary metabolite biosynthesis and stress responses.

📚 Additional Documentation

Notes

(NaAO2_candidate_AO_0-notes.md)

NaAO2_candidate_AO_0 Notes

UniProt curates A0A1J6KBX2 as an L-aspartate oxidase/NadB-family flavoprotein that catalyzes oxidation of L-aspartate to iminoaspartate, linking this candidate to the upstream NAD branch that feeds pyridine alkaloid biosynthesis. [file:NICAT/NaAO2_candidate_AO_0/NaAO2_candidate_AO_0-uniprot.txt "DE RecName: Full=L-aspartate oxidase"; "CC -!- CATALYTIC ACTIVITY: Reaction=L-aspartate + O2 = iminosuccinate + H2O2"; "CC -!- PATHWAY: Alkaloid biosynthesis."; "CC -!- PATHWAY: Cofactor biosynthesis; NAD(+) biosynthesis; iminoaspartate from L-aspartate (oxidase route): step 1/1."]
UniProt also places this candidate in plastid/chloroplast and in the NadB subfamily, which supports conserved primary-metabolism chemistry but does not by itself prove that AO_0 is the pathway-specialized nicotine paralog. [file:NICAT/NaAO2_candidate_AO_0/NaAO2_candidate_AO_0-uniprot.txt "CC -!- SUBCELLULAR LOCATION: Plastid, chloroplast"; "CC -!- SIMILARITY: Belongs to the FAD-dependent oxidoreductase 2 family. NadB subfamily."]
The N. attenuata genome paper argues that nicotine biosynthesis arose through stepwise duplications of ancient NAD and polyamine pathways, so multiple AO-like paralogs are expected and must be resolved explicitly rather than all being treated as core nicotine genes. PMID:28536194
The full glucosylation preprint keeps AO-like chemistry in scope by placing AO2B among the stronger tobacco genes co-expressed with A622, but this is coexpression support rather than direct catalytic assignment and therefore does not resolve AO_0 versus AO_1 in NICAT. [file:projects/NICOTINE_BIOSYNTHESIS/biorxiv-nicotine-glucosylation-notes.md "genes most correlated with A622 include MATE1, PMT3, PMT2, PMT1, beta-GD1, AO2B, and MPO1"; "QPT2 and AO2 are supported at the level of coordinated pathway expression and pathway framing, but this paper does not directly assay their enzymatic roles."]
AO_0 is therefore a plausible NaAO2 candidate, but the current public record does not distinguish it cleanly from AO_1 as the nicotine-pathway-specialized copy. [file:NICAT/NaAO2_candidate_AO_0/NaAO2_candidate_AO_0-uniprot.txt "GN Name=AO_0"; "CC -!- PATHWAY: Alkaloid biosynthesis."]

📄 View Raw YAML

id: A0A1J6KBX2
gene_symbol: NaAO2_candidate_AO_0
product_type: PROTEIN
status: DRAFT
aliases:
- AO_0
- NaAO2
taxon:
  id: NCBITaxon:49451
  label: Nicotiana attenuata
description: >-
  NaAO2_candidate_AO_0 is one of the two current NICAT aspartate oxidase
  candidates for the duplicated pyridine branch feeding nicotine biosynthesis. It
  clearly encodes an L-aspartate oxidase in chloroplast-localized NAD precursor
  metabolism, but the available evidence does not yet distinguish AO_0 from AO_1
  as the specialized nicotine-pathway paralog.
references:
- id: GO_REF:0000002
  title: Gene Ontology annotation through association of InterPro records with GO terms
  findings: []
- id: GO_REF:0000044
  title: Gene Ontology annotation based on UniProtKB/Swiss-Prot Subcellular Location vocabulary mapping, accompanied by conservative changes to GO terms applied by UniProt
  findings: []
- id: GO_REF:0000120
  title: Combined Automated Annotation using Multiple IEA Methods
  findings: []
- id: file:NICAT/NaAO2_candidate_AO_0/NaAO2_candidate_AO_0-uniprot.txt
  title: UniProt entry A0A1J6KBX2 for Nicotiana attenuata AO_0
  findings:
  - statement: AO_0 is annotated as an L-aspartate oxidase in NAD precursor metabolism
    supporting_text: 'DE   RecName: Full=L-aspartate oxidase'
    reference_section_type: DATABASE_ENTRY
  - statement: UniProt places AO_0 in chloroplast-localized NAD biosynthetic chemistry
    supporting_text: 'CC   -!- SUBCELLULAR LOCATION: Plastid, chloroplast'
    reference_section_type: DATABASE_ENTRY
- id: file:NICAT/NaAO2_candidate_AO_0/NaAO2_candidate_AO_0-notes.md
  title: NaAO2 AO_0 candidate notes
  findings:
  - statement: AO_0 is a plausible upstream pyridine-branch nicotine candidate
    supporting_text: UniProt curates A0A1J6KBX2 as an L-aspartate oxidase/NadB-family flavoprotein that catalyzes oxidation of L-aspartate to iminoaspartate, linking this candidate to the upstream NAD branch that feeds pyridine alkaloid biosynthesis.
    reference_section_type: LITERATURE_REVIEW
  - statement: AO_0 versus AO_1 remains an explicit paralog-resolution problem
    supporting_text: AO_0 is therefore a plausible NaAO2 candidate, but the current public record does not distinguish it cleanly from AO_1 as the nicotine-pathway-specialized copy.
    reference_section_type: LITERATURE_REVIEW
existing_annotations:
- term:
    id: GO:0008734
    label: L-aspartate oxidase activity
  evidence_type: IEA
  original_reference_id: GO_REF:0000120
  review:
    summary: This is the specific catalytic activity supported for AO_0.
    action: ACCEPT
    reason: >-
      UniProt and family assignment consistently identify AO_0 as an L-aspartate
      oxidase rather than a generic oxidoreductase.
    supported_by:
    - reference_id: file:NICAT/NaAO2_candidate_AO_0/NaAO2_candidate_AO_0-notes.md
      supporting_text: UniProt curates A0A1J6KBX2 as an L-aspartate oxidase/NadB-family flavoprotein that catalyzes oxidation of L-aspartate to iminoaspartate, linking this candidate to the upstream NAD branch that feeds pyridine alkaloid biosynthesis.
      reference_section_type: LITERATURE_REVIEW
- term:
    id: GO:0009435
    label: NAD+ biosynthetic process
  evidence_type: IEA
  original_reference_id: GO_REF:0000120
  review:
    summary: AO_0 belongs in NAD biosynthesis.
    action: ACCEPT
    reason: >-
      Aspartate oxidase catalyzes the upstream reaction that feeds de novo NAD
      biosynthesis, which is the ancestral source pathway for the pyridine branch.
    supported_by:
    - reference_id: file:NICAT/NaAO2_candidate_AO_0/NaAO2_candidate_AO_0-notes.md
      supporting_text: UniProt curates A0A1J6KBX2 as an L-aspartate oxidase/NadB-family flavoprotein that catalyzes oxidation of L-aspartate to iminoaspartate, linking this candidate to the upstream NAD branch that feeds pyridine alkaloid biosynthesis.
      reference_section_type: LITERATURE_REVIEW
- term:
    id: GO:0009507
    label: chloroplast
  evidence_type: IEA
  original_reference_id: GO_REF:0000044
  review:
    summary: Chloroplast localization is plausible and should be retained as non-core context.
    action: KEEP_AS_NON_CORE
    reason: >-
      The location is supported in UniProt, but the main curation question for
      this candidate is paralog assignment within the pathway rather than
      subcellular detail.
    supported_by:
    - reference_id: file:NICAT/NaAO2_candidate_AO_0/NaAO2_candidate_AO_0-uniprot.txt
      supporting_text: 'CC   -!- SUBCELLULAR LOCATION: Plastid, chloroplast'
      reference_section_type: DATABASE_ENTRY
- term:
    id: GO:0016491
    label: oxidoreductase activity
  evidence_type: IEA
  original_reference_id: GO_REF:0000002
  review:
    summary: This parent oxidoreductase term is true but not specific enough.
    action: MARK_AS_OVER_ANNOTATED
    reason: >-
      The more informative catalytic term GO:0008734 already captures the actual
      chemistry of AO_0.
    supported_by:
    - reference_id: file:NICAT/NaAO2_candidate_AO_0/NaAO2_candidate_AO_0-notes.md
      supporting_text: UniProt curates A0A1J6KBX2 as an L-aspartate oxidase/NadB-family flavoprotein that catalyzes oxidation of L-aspartate to iminoaspartate, linking this candidate to the upstream NAD branch that feeds pyridine alkaloid biosynthesis.
      reference_section_type: LITERATURE_REVIEW
core_functions:
- molecular_function:
    id: GO:0008734
    label: L-aspartate oxidase activity
  directly_involved_in:
  - id: GO:0009435
    label: NAD+ biosynthetic process
  description: >-
    AO_0 is an L-aspartate oxidase candidate in the duplicated NAD-derived branch
    that supplies pyridine precursor chemistry to nicotine biosynthesis.
  supported_by:
  - reference_id: file:NICAT/NaAO2_candidate_AO_0/NaAO2_candidate_AO_0-notes.md
    supporting_text: AO_0 is therefore a plausible NaAO2 candidate, but the current public record does not distinguish it cleanly from AO_1 as the nicotine-pathway-specialized copy.
    reference_section_type: LITERATURE_REVIEW
proposed_new_terms: []
suggested_questions:
- question: Which of the AO_0 and AO_1 paralogs is preferentially induced in nicotine-producing roots after topping or herbivory?
- question: Does only one AO paralog measurably feed the pyridine branch used for nicotine accumulation?
suggested_experiments:
- description: Compare AO_0 and AO_1 transcript abundance across roots, leaves, and induction time courses relevant to nicotine biosynthesis.
  experiment_type: expression profiling
  hypothesis: One AO paralog is preferentially associated with nicotine-producing root tissue.
- description: Create single-paralog knockouts and measure NAD intermediates, nicotinic acid branch metabolites, and nicotine accumulation.
  experiment_type: genetic perturbation plus metabolite profiling
  hypothesis: The pathway-specialized AO paralog will have a stronger effect on nicotine precursor supply.

NaAO2_candidate_AO_0

Gene Description