id: G1FNI6 gene_symbol: TFP product_type: PROTEIN status: COMPLETE taxon: id: NCBITaxon:13288 label: Thlaspi arvense description: TaTFP is a thiocyanate-forming protein of field pennycress, an iron(II)-dependent specifier protein in the glucosinolate-myrosinase chemical defense system. It is a Kelch-repeat, six-bladed beta-propeller protein that acts as a homodimer. Rather than hydrolyzing intact glucosinolates, it captures the unstable aglucone generated by myrosinase upon tissue damage and redirects its rearrangement away from the default isothiocyanate, catalyzing formation of organic thiocyanates (e.g. allylthiocyanate), epithionitriles, and simple nitriles. The protein is curated as a carbon-sulfur lyase (sulfolyase, EC 4.8.1.8) and is expressed constitutively across plant tissues. alternative_records: - id: A0AAU9R9U8 relationship: REFERENCE_PROTEOME_EQUIVALENT reference_proteome: UP000836841 reviewed_on: '2026-10-19' reason: Retain G1FNI6 as the primary Swiss-Prot accession. The leading reference sequence has 97.13% global identity, full target coverage and a short terminal difference, and is separated from other specifier-family hits. This supports a qualified sequence-based counterpart. Its generic epithiospecifier annotation does not independently establish thiocyanate-forming specificity, so functional annotation transfer still requires review. evidence: - type: PROTEOME_SEARCH description: MMseqs2 searched 26,379 sequences across UP000836841 at E <= 1e-5 and sensitivity 7.5, returning 28 hits. Every returned sequence was globally aligned using BLOSUM62 with gap-open -10 and extension -0.5. Scores and sequences for all 28 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:THLAR/TFP/TFP-record-evidence.yaml supporting_text: 'target_sequence_count: 26379' - reference_id: file:THLAR/TFP/TFP-record-evidence.yaml supporting_text: 'candidate_count: 28' - type: SEQUENCE_COMPARISON description: 'A0AAU9R9U8: 338 identities across 348 global-alignment columns (97.1264% identity including gaps), 2 substitutions and 8 gap columns. Query coverage is 97.7011%; target coverage is 100.0%. Sequence similarity is interpreted in the manual rationale. Identity over paired residues is 99.4118% (338/340); gap columns are excluded from this denominator.' supported_by: - reference_id: file:THLAR/TFP/TFP-record-evidence.yaml supporting_text: "query_length: 348\n target_length: 340\n alignment_columns: 348\n identities:\ \ 338\n mismatches: 2\n gap_columns: 8\n identity_percent: 97.1264\n query_coverage_percent:\ \ 97.7011\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 A0AAU9R9U8 in UP000836841 and give it the same taxon as the primary. supported_by: - reference_id: file:THLAR/TFP/TFP-record-evidence.yaml supporting_text: 'primary_in_any_reference_proteome: false' - reference_id: file:THLAR/TFP/TFP-record-evidence.yaml supporting_text: 'alternative_in_reference_proteome: true' - reference_id: file:THLAR/TFP/TFP-record-evidence.yaml supporting_text: 'same_taxon: true' - type: RECORD_HISTORY description: 'Sampled UniSave observations: 11-NOV-2015, entry version 12.: no Proteomes cross-reference; 11-DEC-2019, entry version 25.: no Proteomes cross-reference; 27-NOV-2024, entry version 36.: 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:THLAR/TFP/TFP-record-evidence.yaml supporting_text: "entry_date: 11-NOV-2015, entry version 12.\n proteomes: []" - reference_id: file:THLAR/TFP/TFP-record-evidence.yaml supporting_text: "entry_date: 11-DEC-2019, entry version 25.\n proteomes: []" - reference_id: file:THLAR/TFP/TFP-record-evidence.yaml supporting_text: "entry_date: 27-NOV-2024, entry version 36.\n proteomes: []" existing_annotations: - term: id: GO:0005634 label: nucleus evidence_type: IEA original_reference_id: GO_REF:0000118 qualifier: located_in review: summary: The propagated nuclear localization of TaTFP is not resolved. action: UNDECIDED supported_by: - reference_id: PMID:26260516 - reference_id: PMID:21783213 reason: Arabidopsis ESP has conditional WRKY53-dependent nuclear recruitment; that observation neither demonstrates nor excludes a nuclear pool of pennycress TFP. Direct TaTFP structural and coupled-enzyme evidence does not establish this compartment. Do not call the original localization disproven by the absence of a known nuclear role. - term: id: GO:0005829 label: cytosol evidence_type: IEA original_reference_id: GO_REF:0000118 qualifier: located_in review: summary: Plausible and consistent with function. Specifier proteins act in the cytosol where myrosinase-released glucosinolate aglucones become available. Direct subcellular localization of TaTFP has not been experimentally established, but the cytosol is the expected site of action. action: ACCEPT supported_by: - reference_id: file:THLAR/TFP/TFP-deep-research-falcon.md supporting_text: broader glucosinolate-system reviews/localization studies discuss cytoplasmic organization for specifier proteins and compartmented myrosinases - term: id: GO:0019760 label: glucosinolate metabolic process evidence_type: IEA original_reference_id: GO_REF:0000117 qualifier: involved_in review: summary: Correct, but the generic metabolic-process parent should be specialized to the catabolic child, consistent with the experimentally supported breakdown role of TaTFP downstream of myrosinase. action: MODIFY proposed_replacement_terms: - id: GO:0019762 label: glucosinolate catabolic process - term: id: GO:0030234 label: enzyme regulator activity evidence_type: IEA original_reference_id: GO_REF:0000118 qualifier: enables review: summary: Outdated molecular-function framing. Specifier proteins were historically viewed as accessory factors that "regulate" myrosinase product outcome, but TaTFP is now established as a catalyst in its own right - an Fe(2+)-dependent carbon-sulfur lyase (sulfolyase, EC 4.8.1.8) that eliminates sulfate from the glucosinolate aglucone to form thiocyanates and nitriles. The activity should be captured as a lyase, not as enzyme regulator activity. [PMID:30900313 "specifier proteins act as Fe(2+)-dependent lyases"] action: MODIFY proposed_replacement_terms: - id: GO:0016846 label: carbon-sulfur lyase activity supported_by: - reference_id: PMID:30900313 supporting_text: 'Structural diversification during glucosinolate breakdown: mechanisms of thiocyanate, epithionitrile and simple nitrile formation' - reference_id: PMID:21783213 supporting_text: A thiocyanate-forming protein generates multiple products upon allylglucosinolate breakdown in Thlaspi arvense. - reference_id: PMID:23999604 supporting_text: Molecular models and mutational analyses of plant specifier proteins suggest active site residues and reaction mechanism - term: id: GO:0080028 label: nitrile biosynthetic process evidence_type: IEA original_reference_id: GO_REF:0000118 qualifier: involved_in review: summary: Correct. TaTFP generates simple nitriles and epithionitriles (e.g. 3,4-epithiobutanenitrile) from glucosinolate aglucones, in addition to organic thiocyanates. [PMID:21783213; PMID:30900313] action: ACCEPT supported_by: - reference_id: PMID:21783213 supporting_text: A thiocyanate-forming protein generates multiple products upon allylglucosinolate breakdown in Thlaspi arvense. - reference_id: PMID:30900313 supporting_text: mechanisms of thiocyanate, epithionitrile and simple nitrile formation - term: id: GO:0019760 label: glucosinolate metabolic process evidence_type: IDA original_reference_id: PMID:26260516 qualifier: involved_in review: summary: Experimentally supported but can be made more precise. TaTFP acts in the breakdown (catabolism) of glucosinolates downstream of myrosinase, so the catabolic child term is more informative than the generic metabolic-process parent. [PMID:26260516 "a kelch protein involved in glucosinolate breakdown"] action: MODIFY proposed_replacement_terms: - id: GO:0019762 label: glucosinolate catabolic process supported_by: - reference_id: PMID:26260516 supporting_text: The crystal structure of the thiocyanate-forming protein from Thlaspi arvense, a kelch protein involved in glucosinolate breakdown - term: id: GO:0042802 label: identical protein binding evidence_type: IDA original_reference_id: PMID:26260516 qualifier: enables review: summary: Reflects the homodimeric quaternary state demonstrated in the crystal structure. Real but structural rather than the catalytic core function; retained as non-core. The homodimerization-activity annotation below is the more specific of the two. action: KEEP_AS_NON_CORE supported_by: - reference_id: PMID:26260516 supporting_text: crystal structure of the thiocyanate-forming protein from Thlaspi arvense - term: id: GO:0042803 label: protein homodimerization activity evidence_type: IDA original_reference_id: PMID:26260516 qualifier: enables review: summary: Supported by the crystal structure, which shows TaTFP is a homodimer. A genuine structural property of the protein, retained as non-core relative to its catalytic lyase activity. [PMID:26260516] action: KEEP_AS_NON_CORE supported_by: - reference_id: PMID:26260516 supporting_text: TaTFP crystallized as homodimer. core_functions: - description: TaTFP is an Fe(2+)-dependent carbon-sulfur lyase (sulfolyase) that captures the myrosinase-generated glucosinolate aglucone and eliminates sulfate to form organic thiocyanates, epithionitriles, and simple nitriles, thereby diversifying glucosinolate breakdown chemistry away from the default isothiocyanate. molecular_function: id: GO:0016846 label: carbon-sulfur lyase activity directly_involved_in: - id: GO:0019762 label: glucosinolate catabolic process - id: GO:0080028 label: nitrile biosynthetic process locations: - id: GO:0005829 label: cytosol supported_by: - reference_id: PMID:21783213 supporting_text: this protein promotes the formation of allylthiocyanate as well as the corresponding epithionitrile upon myrosinase-catalyzed hydrolysis of allylglucosinolate, the major glucosinolate of T. arvense. - reference_id: PMID:30900313 supporting_text: Based on these insights, we propose that specifier proteins are catalysts that might be classified as Fe2+ -dependent lyases. - reference_id: PMID:26260516 supporting_text: TaTFP crystallized as homodimer. Each monomer forms a six-blade β-propeller with a wide "top" and a narrower "bottom" opening with distinct strand-connecting loops protruding far beyond the lower propeller surface. references: - id: file:THLAR/TFP/TFP-record-evidence.yaml title: Compact archived record and sequence evidence for G1FNI6 - id: GO_REF:0000117 title: Electronic Gene Ontology annotations created by ARBA machine learning models findings: [] - id: GO_REF:0000118 title: TreeGrafter-generated GO annotations findings: [] - id: PMID:26260516 title: The crystal structure of the thiocyanate-forming protein from Thlaspi arvense, a kelch protein involved in glucosinolate breakdown. findings: - statement: TaTFP is a Kelch-repeat six-bladed beta-propeller protein that forms a homodimer and functions in glucosinolate breakdown. reference_section_type: RESULTS reference_review: relevance: HIGH correctness: VERIFIED review_notes: Source of the IDA annotations for glucosinolate breakdown, identical-protein binding, and homodimerization; PDB 5A10/5A11. - id: PMID:21783213 title: A thiocyanate-forming protein generates multiple products upon allylglucosinolate breakdown in Thlaspi arvense. findings: - statement: TaTFP catalyzes allylthiocyanate and epithionitrile formation from allylglucosinolate in the presence of myrosinase, is Fe(2+)-stimulated and EDTA-repressed, and is expressed constitutively across tissues. reference_section_type: RESULTS reference_review: relevance: HIGH correctness: VERIFIED review_notes: Primary biochemical characterization; basis for EC 4.8.1.8 and the Fe(2+) cofactor requirement. Abstract-only in cache. - id: PMID:23999604 title: Molecular models and mutational analyses of plant specifier proteins suggest active site residues and reaction mechanism. findings: - statement: Homology models and mutagenesis of specifier proteins implicate active-site residues governing product specificity. reference_section_type: RESULTS reference_review: relevance: MEDIUM correctness: VERIFIED review_notes: Supports mechanistic/active-site interpretation. Abstract-only in cache. - id: PMID:30900313 title: 'Structural diversification during glucosinolate breakdown: mechanisms of thiocyanate, epithionitrile and simple nitrile formation.' findings: - statement: Specifier proteins, including TaTFP, are reclassified as Fe(2+)-dependent lyases; the iron-binding triad (E266/D270/H274) and substrate docking pose determine whether thiocyanate, epithionitrile, or simple nitrile is produced. reference_section_type: RESULTS reference_review: relevance: HIGH correctness: VERIFIED review_notes: Establishes the lyase (catalyst) classification underpinning the MODIFY of enzyme regulator activity to carbon-sulfur lyase activity. Full text available. - id: file:THLAR/TFP/TFP-deep-research-falcon.md title: Falcon/Edison deep research report for TFP (TaTFP), Thlaspi arvense findings: - statement: Synthesizes TaTFP as an Fe(2+)-dependent Kelch beta-propeller specifier protein in glucosinolate breakdown; flags that direct subcellular localization of TaTFP is not experimentally established. reference_section_type: OTHER proposed_new_terms: - proposed_name: thiocyanate-forming sulfolyase activity proposed_definition: A carbon-sulfur lyase activity that eliminates sulfate from a glucosinolate aglucone to yield an organic thiocyanate (EC 4.8.1.8). No GO molecular-function term currently captures this specific specifier-protein activity; carbon-sulfur lyase activity (GO:0016846) is the closest available parent. justification: The reaction is curated in UniProt (EC 4.8.1.8) and structurally characterized, but the existing GO MF granularity stops at the carbon-sulfur lyase parent, leaving specifier-protein catalysis under-described. proposed_parent: id: GO:0016846 label: carbon-sulfur lyase activity suggested_questions: - question: What is the steady-state subcellular localization of TaTFP in intact pennycress tissue, and how does it co-localize with myrosinase prior to tissue disruption? suggested_experiments: - description: Determine TaTFP subcellular localization by fluorescent-protein fusion or immunolocalization in Thlaspi arvense, and test co-compartmentalization with myrosinase to define where aglucone capture occurs in vivo.