id: Q945B5 gene_symbol: AOP2 product_type: PROTEIN status: COMPLETE taxon: id: NCBITaxon:3702 label: Arabidopsis thaliana description: AOP2 (ALKENYL HYDROXALKYL PRODUCING 2; At4g03060) is an Fe(II)- and 2-oxoglutarate-dependent dioxygenase (2-ODD) that catalyzes a side-chain modification in methionine-derived (aliphatic) glucosinolate biosynthesis, converting methylsulfinylalkyl glucosinolates into alkenyl glucosinolates (e.g. 3-methylsulfinylpropyl to allyl/2-propenyl glucosinolate, and 4-methylsulfinylbutyl glucoraphanin to 3-butenyl glucosinolate). It underlies the GS-ALK locus. AOP2 arose by tandem duplication with AOP3, which instead forms hydroxyalkyl glucosinolates, and the two are expressed in a mutually exclusive manner across accessions. This entry is the functional allele from the Cape Verde Islands (Cvi) accession (432 aa, AOP2-1); the Columbia allele carries a 5-bp deletion causing a frameshift and a non-functional truncated protein, so Col-0 accumulates methylsulfinylalkyl glucosinolates. AOP2 is expressed most strongly in leaves and stems in a light-dependent manner, and its activity shapes the glucosinolate profile and thus the isothiocyanate breakdown products that mediate defense against herbivores. alternative_records: - id: Q9ZTA2 relationship: REFERENCE_PROTEOME_LOCUS_COUNTERPART reference_proteome: UP000006548 reviewed_on: '2026-10-19' reason: Retain Q945B5 as the primary Swiss-Prot accession. UniProt explicitly identifies Q9ZTA2 as the Columbia AOP2 allele with a 5-bp deletion leading to a non-functional protein, contrasting the functional Cvi allele Q945B5. The observed sequence differences and length loss are consistent with that curated distinction. Record same-locus correspondence, not equivalent catalytic function. evidence: - type: PROTEOME_SEARCH description: MMseqs2 searched 39,272 sequences across UP000006548 at E <= 1e-5 and sensitivity 7.5, returning 106 hits. Every returned sequence was globally aligned using BLOSUM62 with gap-open -10 and extension -0.5. Scores and sequences for all 106 returned candidates are retained, with 10 detailed alignment files. The heuristic search does not establish genomic absence or unique locus identity. supported_by: - reference_id: file:ARATH/AOP2/AOP2-record-evidence.yaml supporting_text: 'target_sequence_count: 39272' - reference_id: file:ARATH/AOP2/AOP2-record-evidence.yaml supporting_text: 'candidate_count: 106' - type: SEQUENCE_COMPARISON description: 'Q9ZTA2: 390 identities across 432 global-alignment columns (90.2778% identity including gaps), 9 substitutions and 33 gap columns. Query coverage is 92.3611%; target coverage is 100.0%. Sequence similarity is interpreted in the manual rationale. Identity over paired residues is 97.7444% (390/399); gap columns are excluded from this denominator.' supported_by: - reference_id: file:ARATH/AOP2/AOP2-record-evidence.yaml supporting_text: "query_length: 432\n target_length: 399\n alignment_columns: 432\n identities:\ \ 390\n mismatches: 9\n gap_columns: 33\n identity_percent: 90.2778\n query_coverage_percent:\ \ 92.3611\n target_coverage_percent: 100.0" - type: PROTEOME_MEMBERSHIP description: The primary accession is outside every searched exact-taxon reference proteome. Current UniProt records place Q9ZTA2 in UP000006548 and give it the same taxon as the primary. supported_by: - reference_id: file:ARATH/AOP2/AOP2-record-evidence.yaml supporting_text: 'primary_in_any_reference_proteome: false' - reference_id: file:ARATH/AOP2/AOP2-record-evidence.yaml supporting_text: 'alternative_in_reference_proteome: true' - reference_id: file:ARATH/AOP2/AOP2-record-evidence.yaml supporting_text: 'same_taxon: true' - type: RECORD_HISTORY description: 'Sampled UniSave observations: 24-JUN-2015, entry version 46.: no Proteomes cross-reference; 26-FEB-2020, entry version 59.: no Proteomes cross-reference; 27-NOV-2024, entry version 68.: no Proteomes cross-reference. These dated observations do not establish uninterrupted membership/absence, the exact change date, or UniProt''s motivation.' supported_by: - reference_id: file:ARATH/AOP2/AOP2-record-evidence.yaml supporting_text: "entry_date: 24-JUN-2015, entry version 46.\n proteomes: []" - reference_id: file:ARATH/AOP2/AOP2-record-evidence.yaml supporting_text: "entry_date: 26-FEB-2020, entry version 59.\n proteomes: []" - reference_id: file:ARATH/AOP2/AOP2-record-evidence.yaml supporting_text: "entry_date: 27-NOV-2024, entry version 68.\n proteomes: []" - type: CURATION_CONTEXT description: UniProt directly identifies the Columbia AOP2 deletion allele and the functional Cvi counterpart. supported_by: - reference_id: file:ARATH/AOP2/AOP2-record-evidence.yaml record_section: - CC:CAUTION supporting_text: 5-bp deletion that leads to a non-functional protein - reference_id: file:ARATH/AOP2/AOP2-record-evidence.yaml record_section: - CC:CAUTION supporting_text: functional and expressed alleles for AOP2 (AC Q945B5) existing_annotations: - term: id: GO:0016706 label: 2-oxoglutarate-dependent dioxygenase activity evidence_type: IDA original_reference_id: PMID:11251105 qualifier: enables review: summary: Heterologously expressed AOP2 (thioredoxin fusion in E. coli), assayed with 2-oxoglutarate and ascorbate, converted 3-methylsulfinylpropyl and 4-methylsulfinylbutyl glucosinolates to allyl and 3-butenyl glucosinolates. action: ACCEPT reason: Correct and experimentally supported. GO has no specific term for the alkenyl glucosinolate-forming reaction, so the 2-oxoglutarate-dependent dioxygenase activity class term is the most specific existing MF term; a more precise new term is proposed (see proposed_new_terms). The assay included 2-oxoglutarate, and the protein contains the conserved Fe(II)/2OG-binding residues of the 2OG-FeII oxygenase domain. supported_by: - reference_id: PMID:11251105 supporting_text: induced AOP2 bacterial extracts catalyzed the conversion of the 3-methylsulfinylpropyl to the allyl glucosinolate and the 4-methylsulfinylbutyl to the 3-butenyl glucosinolate - reference_id: PMID:11251105 supporting_text: with the addition of 200 mM sucrose and the use of 10 mM oxoglutarate and 10 mM ascorbate instead of 40 mM of each of these - term: id: GO:0016706 label: 2-oxoglutarate-dependent dioxygenase activity evidence_type: IEA original_reference_id: GO_REF:0000117 qualifier: enables supporting_entities: - ARBA:ARBA00027633 review: summary: ARBA machine-learning rule assigns 2-oxoglutarate-dependent dioxygenase activity based on the 2OG-FeII oxygenase domain. action: ACCEPT reason: Consistent with the experimental IDA annotation and the domain architecture (DIOX_N and 2OG-FeII_Oxy Pfam domains; Fe-binding His/Asp and 2OG-binding Arg residues). supported_by: - reference_id: PMID:11251105 supporting_text: we report the identification of genes encoding two 2-oxoglutarate--dependent dioxygenases that are responsible for this variation - term: id: GO:0019761 label: glucosinolate biosynthetic process evidence_type: IDA original_reference_id: PMID:11251105 qualifier: involved_in review: summary: AOP2 catalyzes the formation of alkenyl glucosinolates from methylsulfinylalkyl glucosinolate precursors, a secondary side-chain modification step of aliphatic glucosinolate biosynthesis. action: ACCEPT reason: AOP2 directly catalyzes a biosynthetic step (enzyme assay), and in planta overexpression of a functional AOP2 allele in Col-0 converts methylsulfinylalkyl glucosinolates to 2-propenyl and 3-butenyl glucosinolates. Core process. supported_by: - reference_id: PMID:11251105 supporting_text: we show that AOP2 catalyzes the conversion of methylsulfinylalkyl glucosinolates to alkenyl glucosinolates - reference_id: PMID:20699011 supporting_text: The over-expression of a functional AOP2 allele was found to successfully convert the precursor methylsulfinyl alkyl glucosinolate into the alkenyl form. - term: id: GO:0019761 label: glucosinolate biosynthetic process evidence_type: IEA original_reference_id: GO_REF:0000117 qualifier: involved_in supporting_entities: - ARBA:ARBA00085760 review: summary: ARBA rule assigns glucosinolate biosynthetic process, consistent with the experimental evidence. action: ACCEPT reason: Matches the experimentally established role of AOP2 in aliphatic glucosinolate side-chain modification. supported_by: - reference_id: PMID:20699011 supporting_text: In Arabidopsis the AOP2 gene plays a role in the secondary modification of aliphatic (methionine-derived) glucosinolates, namely the conversion of methylsulfinylalkyl glucosinolates to form alkenyl glucosinolates core_functions: - description: The functional AOP2 allele represented by Q945B5 catalyzes Fe(II)/2-oxoglutarate-dependent conversion of methylsulfinylalkyl glucosinolates to alkenyl glucosinolates, including allyl and 3-butenyl products. This activity is not attributed to the frameshifted, nonfunctional endogenous Col-0 allele. proposed_molecular_function: alkenyl glucosinolate synthase activity directly_involved_in: - id: GO:0019761 label: glucosinolate biosynthetic process supported_by: - reference_id: PMID:11251105 supporting_text: This finding shows that AOP2 is capable of catalyzing the GS-ALK reaction with both the 3-methylsulfinylalkyl and 4-methylsulfinylalkyl glucosinolates. - reference_id: PMID:20699011 supporting_text: showed that the AOP2-2 allele catalysed the conversion of the two precursor methylsulfinylalkyl glucosinolates to 2-propenyl and 3-butenyl glucosinolates proposed_new_terms: - proposed_name: alkenyl glucosinolate synthase activity proposed_definition: Catalysis of the Fe(II)- and 2-oxoglutarate-dependent conversion of an omega-(methylsulfinyl)alkyl glucosinolate to the corresponding omega-alkenyl glucosinolate with loss of the methylsulfinyl group, e.g. 3-methylsulfinylpropyl glucosinolate to allyl (2-propenyl) glucosinolate, or 4-methylsulfinylbutyl glucosinolate (glucoraphanin) to 3-butenyl glucosinolate (gluconapin). justification: AOP2 (GS-ALK) has a biochemically demonstrated, substrate-defined activity that determines alkenyl glucosinolate accumulation across Arabidopsis accessions and Brassica crops (BoGSL-ALK), but GO only offers the generic class term GO:0016706. The paralog AOP3 forms hydroxyalkyl glucosinolates from the same substrates, so a specific term would distinguish these neofunctionalized duplicates. Should be placed under 2-oxoglutarate-dependent dioxygenase activity (GO:0016706). proposed_parent: id: GO:0016706 label: 2-oxoglutarate-dependent dioxygenase activity supported_by: - reference_id: PMID:11251105 supporting_text: induced AOP2 bacterial extracts catalyzed the conversion of the 3-methylsulfinylpropyl to the allyl glucosinolate and the 4-methylsulfinylbutyl to the 3-butenyl glucosinolate - reference_id: PMID:20699011 supporting_text: The AOP2 homologue (BoGSL-ALK) from collard (Brassica oleracea) has been cloned and characterised and it was found that the gene product is able to catalyse the conversion of methylsulfinylalkyl glucosinolate to the alkenyl form in planta references: - id: file:ARATH/AOP2/AOP2-record-evidence.yaml title: Compact archived record and sequence evidence for Q945B5 - id: GO_REF:0000117 title: Electronic Gene Ontology annotations created by ARBA machine learning models findings: - statement: ARBA rules assign 2-oxoglutarate-dependent dioxygenase activity and glucosinolate biosynthetic process, both consistent with experimental evidence. - id: PMID:11251105 title: 'Gene duplication in the diversification of secondary metabolism: tandem 2-oxoglutarate-dependent dioxygenases control glucosinolate biosynthesis in Arabidopsis.' findings: - statement: Identifies AOP2 and AOP3 as tandem 2-ODDs at the GS-AOP locus; AOP2 expressed in E. coli converts 3-methylsulfinylpropyl and 4-methylsulfinylbutyl glucosinolates to allyl and 3-butenyl glucosinolates; Col-0 carries a frameshifted non-functional AOP2; no ecotype co-expresses AOP2 and AOP3. supporting_text: we show that AOP2 catalyzes the conversion of methylsulfinylalkyl glucosinolates to alkenyl glucosinolates reference_section_type: ABSTRACT reference_review: relevance: HIGH correctness: VERIFIED review_notes: Full text cached; source of IDA MF and BP annotations. - id: PMID:20699011 title: 'The characterisation of AOP2: a gene associated with the biosynthesis of aliphatic alkenyl glucosinolates in Arabidopsis thaliana.' findings: - statement: Defines three AOP2 alleles (AOP2-1 Cvi/St, AOP2-2 Pi, AOP2-3 Col/Ler with 5-bp frameshift); AOP2 is light-regulated and most expressed in leaf and stem; overexpression of AOP2-2 in Col-0 converts methylsulfinylalkyl glucosinolates to 2-propenyl and 3-butenyl glucosinolates in planta. supporting_text: The over-expression of a functional AOP2 allele was found to successfully convert the precursor methylsulfinyl alkyl glucosinolate into the alkenyl form. reference_section_type: ABSTRACT reference_review: relevance: HIGH correctness: VERIFIED review_notes: Full text cached; in planta confirmation of AOP2 activity. - id: PMID:25758208 title: The Glucosinolate Biosynthetic Gene AOP2 Mediates Feed-back Regulation of Jasmonic Acid Signaling in Arabidopsis. findings: - statement: AOP2 participates in a feedback loop linking glucosinolate biosynthesis to JA signaling, partly independent of MYB28/MYB29; mechanism unresolved, so no GO process term is proposed. supporting_text: Transcriptome analysis of these mutants identified a so far unrecognized link between AOP2 and jasmonic acid (JA) signaling independent of MYB28 and MYB29. reference_review: relevance: MEDIUM correctness: VERIFIED review_notes: Cached identifier/title and the relevant abstract or available full text checked during the module-member audit. The scope of target-specific evidence is qualified in annotation reasons and notes; a cached abstract is not treated as full text. - id: PMID:26442014 title: Natural variation in cross-talk between glucosinolates and onset of flowering in Arabidopsis. findings: - statement: AOP2 introduced into AOP-null accessions alters glucosinolate levels and, depending on genetic background, flowering time. supporting_text: the effect on flowering time was dependent on the genetic background pointing to natural variation in cross-talk between defense chemistry and onset of flowering reference_review: relevance: MEDIUM correctness: VERIFIED review_notes: Cached identifier/title and the relevant abstract or available full text checked during the module-member audit. The scope of target-specific evidence is qualified in annotation reasons and notes; a cached abstract is not treated as full text. - id: PMID:20303821 title: Biosynthesis of glucosinolates--gene discovery and beyond. findings: - statement: Review placing AOP2 in the side-chain decoration phase of glucosinolate biosynthesis. supporting_text: Recent progress includes genes in all three phases of the pathway, i.e. side-chain elongation of precursor amino acids, formation of the core glucosinolate structure and side-chain decoration. reference_section_type: ABSTRACT reference_review: relevance: LOW correctness: VERIFIED review_notes: Abstract-only review, used for pathway context. suggested_questions: - question: What is the chemical mechanism and by-product of AOP2-catalyzed alkenyl formation (loss of methanesulfenic acid?), and does AOP2 retain the canonical hydroxylation chemistry of 2-ODDs as a cryptic activity? - question: Which active-site residues distinguish AOP2 (alkenyl-forming) from AOP3 (hydroxyalkyl-forming), given that the two arose by recent tandem duplication? - question: AOP2 expression raises MYB28/MYB29 transcripts and alters jasmonate signaling independently of MYB28/MYB29 (PMID:25758208), and AOP2 alters flowering time in a background-dependent way (PMID:26442014). Are these effects mediated by alkenyl glucosinolates or their breakdown products, or by the AOP2 protein itself? suggested_experiments: - description: Purify recombinant AOP2 and AOP3 and perform steady-state kinetics with C3-C8 methylsulfinylalkyl glucosinolates, including product and by-product identification by LC-MS, to define specificity and mechanism. - description: Swap residues between AOP2 and AOP3 guided by homology models or crystal structures to identify determinants of alkenylation versus hydroxylation. - description: Complement a Col-0 background (non-functional AOP2) with catalytically dead AOP2 (Fe-binding His mutants) versus wild-type AOP2 to separate enzymatic from possible non-enzymatic effects on glucosinolate accumulation and downstream traits.