{"gene":"METAP1","run_date":"2026-06-10T02:59:50","timeline":{"discoveries":[{"year":1995,"finding":"S. cerevisiae MetAP1 (Map1p) is a methionine aminopeptidase that cotranslationally removes N-terminal methionine from nascent polypeptides; it contains an N-terminal zinc-finger domain absent in prokaryotic homologs. Deletion of MAP1 is viable but causes slow growth; the double map1/map2 null is nonviable, establishing that N-terminal methionine removal is an essential function requiring two MetAPs.","method":"Genetic deletion, complementation assay, immunoaffinity purification, enzymatic activity assay","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — genetic epistasis (double null lethality), biochemical purification and enzymatic assay, replicated across multiple labs subsequently","pmids":["8618900"],"is_preprint":false},{"year":1997,"finding":"Fumagillin selectively inhibits S. cerevisiae MetAP2 in vivo but does NOT inhibit MetAP1 in vivo; MetAP1 function is insufficient to substitute for MetAP2 when MetAP2 is covalently blocked by fumagillin.","method":"In vivo yeast growth assay with fumagillin, differential inhibition of MetAP1 vs MetAP2","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo yeast genetic and pharmacological experiments, replicated across subsequent studies","pmids":["9177176"],"is_preprint":false},{"year":1997,"finding":"Mutation of the conserved cobalt-coordinating residue Asp219 (analogous to Asp97 in E. coli MetAP) in yeast MetAP1 to Asn reduces catalytic activity ~1000-fold and alters substrate specificity, demonstrating that Asp219 is essential for catalysis. The D219N mutant acts as a dominant negative, interfering with both wild-type MetAP1 and MetAP2 function in yeast.","method":"Site-directed mutagenesis, enzymatic activity assay, in vivo yeast growth assay","journal":"Archives of biochemistry and biophysics","confidence":"High","confidence_rationale":"Tier 1 / Moderate — active-site mutagenesis combined with in vitro enzymatic assay and in vivo dominant-negative phenotype, single lab but multiple orthogonal methods","pmids":["9367524"],"is_preprint":false},{"year":2002,"finding":"Yeast MetAP1 is a ribosome-associated protein that primarily associates with the 60S ribosomal subunit and 80S translational complex. The N-terminal zinc finger domain is required for this association: single point mutations in the first or second zinc finger motif disrupt association with the 60S subunit and 80S complex and reduce N-terminal methionine removal from a reporter protein from ~100% to 31–35%.","method":"Ribosome sedimentation profiling, zinc finger mutagenesis, reporter protein N-terminal processing assay","journal":"Journal of cellular biochemistry","confidence":"High","confidence_rationale":"Tier 2 / Moderate — ribosome fractionation plus mutagenesis plus functional processing assay, multiple orthogonal methods in one study","pmids":["11968008"],"is_preprint":false},{"year":2002,"finding":"MetAP1 plays the dominant role in N-terminal methionine removal in S. cerevisiae in vivo. Both MetAP1 and MetAP2 are less efficient when the penultimate residue is Val; MetAP2 is less efficient than MetAP1 for Gly, Cys, or Thr at this position, establishing different in vivo cleavage specificities.","method":"In vivo N-terminal processing assay of mutant GST reporter proteins in map1, map2, and wild-type yeast strains","journal":"Archives of biochemistry and biophysics","confidence":"High","confidence_rationale":"Tier 2 / Moderate — systematic in vivo substrate specificity profiling with multiple deletion strains and multiple substrates","pmids":["11811952"],"is_preprint":false},{"year":2002,"finding":"Human MetAP1 functionally complements yeast map1 null growth defect, demonstrating conserved function between yeast and human MetAP1 in vivo.","method":"Heterologous expression of human MetAP1 in yeast map1 null strain, complementation growth assay","journal":"Protein and peptide letters","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — single lab, single complementation method","pmids":["12144506"],"is_preprint":false},{"year":2003,"finding":"Yeast MetAP1 plays a significant role in methionine salvage metabolism, preventing premature activation of MET genes. In cells lacking MetAP1, excess methionine causes growth inhibition by product inhibition of MetAP2 (not MetAP1), revealing differential regulation and a functional distinction between the two isoforms.","method":"Genetic deletion strains, MET gene reporter assays, growth inhibition assays with methionine supplementation","journal":"Journal of cellular biochemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple genetic and metabolic assays in single lab","pmids":["12874831"],"is_preprint":false},{"year":2003,"finding":"A single active-site residue difference between MetAP1 and MetAP2 accounts for differential sensitivity to ovalicin: the analogous position to MetAP2 Ala362 in MetAP1 is naturally a threonine, conferring resistance. Mutating MetAP1 Thr to Ala renders MetAP1 ovalicin-sensitive, and mutating MetAP2 Ala362 to Thr confers ovalicin resistance.","method":"Yeast-based mutagenesis screen, site-directed mutagenesis, in vivo inhibitor sensitivity assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — bidirectional mutagenesis (MetAP1 and MetAP2) with functional validation in yeast, defines molecular basis of specificity","pmids":["14676204"],"is_preprint":false},{"year":2005,"finding":"Crystal structure of human MetAP1 reveals that its active site is smaller than that of human MetAP2, explaining why ovalicin/fumagillin preferentially targets MetAP2. The N-terminal region of MetAP1 contains three Pro-x-x-Pro motifs consistent with ribosome binding.","method":"X-ray crystallography, structural comparison with MetAP2–ovalicin complex","journal":"Biochemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — crystal structure at defined resolution with comparative structural analysis explaining functional differences","pmids":["16274222"],"is_preprint":false},{"year":2005,"finding":"siRNA-mediated knockdown of MetAP1 significantly inhibits proliferation of human umbilical vein endothelial cells (HUVEC, 70–80% inhibition) and A549 lung carcinoma cells (20–30%). Combined knockdown of MetAP1 and MetAP2 produces near-complete growth inhibition, mirroring the map1/map2 double null yeast phenotype.","method":"siRNA knockdown, cell proliferation assay","journal":"Journal of cellular biochemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — siRNA knockdown with quantitative proliferation readout, multiple cell lines tested","pmids":["15962312"],"is_preprint":false},{"year":2006,"finding":"Ovalicin forms a low-affinity covalent adduct with the active-site histidine of human MetAP1 (His310), analogous to the covalent modification of His231 in MetAP2, but with different inhibitor alignment. Several active-site residues must shift outward to accommodate the inhibitor, explaining the lower affinity for MetAP1.","method":"X-ray crystallography at 1.1 Å resolution of ovalicin–MetAP1 complex","journal":"Protein science","confidence":"High","confidence_rationale":"Tier 1 / Moderate — high-resolution crystal structure of inhibitor–enzyme complex with structural comparison","pmids":["16823043"],"is_preprint":false},{"year":2006,"finding":"Human MetAP1 (HsMetAP1) is required for normal G2/M phase cell cycle progression. Selective MetAP1 inhibitors (pyridine-2-carboxylic acid class) cause G2/M accumulation in tumor cells, induce apoptosis in leukemia lines, and cause N-terminal methionine retention in a known MetAP substrate. Overexpression of HsMetAP1 (but not MetAP2) confers resistance, and siRNA knockdown of HsMetAP1 recapitulates slow G2/M progression.","method":"Enzymatic assay, X-ray crystallography, cell cycle FACS analysis, siRNA knockdown, overexpression rescue, N-terminal processing assay","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — multiple orthogonal methods (structure, enzymatic assay, siRNA, overexpression rescue, cell cycle analysis) in single study","pmids":["17114291"],"is_preprint":false},{"year":2007,"finding":"Human cytosolic MetAP1 requires at least three Co2+ ions for optimal catalysis (Hill coefficient n≈2.9). The conserved residue His212 coordinates a third Co2+ ion unique to this enzyme; H212A and H212K mutations reduce kcat 60- and 1800-fold respectively, increase K0.5 for Co2+, and decrease cooperativity, establishing that three metal ions are functionally required—more than any other MetAP family member.","method":"Kinetic analysis, site-directed mutagenesis, Co2+-activation curves, in vitro enzymatic assay","journal":"Biochemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — rigorous kinetic analysis combined with active-site mutagenesis of two independent mutations, multiple kinetic parameters measured","pmids":["17929833"],"is_preprint":false},{"year":2010,"finding":"Systematic profiling of human MetAP1 substrate specificity using a combinatorial peptide library and kinetic analysis reveals: MetAP1 requires small residues (Gly, Ala, Ser, Cys, Pro, Thr, Val) at P1'; has lower activity toward Met-Val and Met-Thr substrates compared with MetAP2; is poorly active toward peptides with Pro at P2'; and disfavors acidic residues at P2'–P5'.","method":"Combinatorial peptide library screening, kinetic analysis of individual peptide substrates","journal":"Biochemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — systematic library screen plus individual kinetic measurements, defines substrate specificity rules","pmids":["20521764"],"is_preprint":false},{"year":2015,"finding":"The N-terminal zinc-binding domain (ZBD, residues 1–83) of human MetAP1 contains two α-helical fragments (residues 44–49 and 59–82) and unfolds upon EDTA chelation of zinc, as demonstrated by NMR chemical shift assignments.","method":"NMR spectroscopy (15N-HSQC, chemical shift assignment), EDTA perturbation","journal":"Biomolecular NMR assignments","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — NMR structural characterization, single lab, provides partial structural information of isolated domain without full structure or functional validation","pmids":["25921012"],"is_preprint":false},{"year":2016,"finding":"Both MetAP1 and MetAP2 are required in vivo for N-terminal methionine excision from M[VT]X-class substrates. Cell sensitivity to fumagillin (MetAP2 inhibitor) correlates with MetAP1 protein levels and with the ability to modulate glutathione homeostasis; fumagillin-sensitive cells show glutathione redox alterations absent in resistant cells.","method":"Large-scale N-terminus proteomics (N-terminomics), fumagillin treatment in multiple cell lines, proteo-transcriptomic analysis","journal":"Oncotarget","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — global N-terminome profiling in multiple cell lines, single lab","pmids":["27542228"],"is_preprint":false},{"year":2022,"finding":"Zng1 (human ortholog conserved) is a GTP-dependent metallochaperone that transfers Zn2+ or Co2+ to apo-MetAP1 (Map1p) in vitro, requiring GTP hydrolysis for metal transfer—unlike known copper chaperones. Deletion of ZNG1 in yeast causes defective Map1p function; Zng1 physically interacts with Map1p.","method":"In vitro metal transfer assay, GTPase assay, yeast deletion genetics, pulldown interaction assay, proteomics","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — in vitro reconstitution of metal transfer, genetic epistasis in yeast, physical interaction demonstrated, multiple orthogonal methods","pmids":["35584675"],"is_preprint":false},{"year":2023,"finding":"In eukaryotes, the nascent polypeptide-associated complex (NAC) controls ribosome binding of METAP1. NAC recruits METAP1 via a long flexible tail and provides a platform for formation of an active methionine excision complex at the ribosomal tunnel exit, ensuring efficient methionine excision from cytosolic proteins while sparing ER-targeted proteins.","method":"Biochemical interaction assays, structural studies (cryo-EM), in vivo functional studies","journal":"Science","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — cryo-EM structure plus biochemical reconstitution plus in vivo validation, mechanistically defines how METAP1 accesses ribosome","pmids":["37347872"],"is_preprint":false},{"year":2023,"finding":"MetAP1 is a novel cisplatin-binding protein: cisplatin binds to MetAP1 via cysteine chelation and contributes to cisplatin's cytotoxicity, as demonstrated by competitive activity-based protein profiling and functional validation.","method":"Competitive activity-based protein profiling (ABPP), functional cytotoxicity validation","journal":"RSC chemical biology","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — competitive ABPP with functional validation, single lab, limited mechanistic depth in abstract","pmids":["37654507"],"is_preprint":false},{"year":2024,"finding":"NAC assembles a multienzyme complex with MetAP1 and NatA early during translation, pre-positioning the active sites of both enzymes for sequential cotranslational processing (methionine excision then N-terminal acetylation) of nascent proteins. NAC also releases inhibitory interactions from the NatA regulatory protein HYPK to activate NatA on the ribosome.","method":"Biochemical assays, cryo-EM structural studies, in vivo studies","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM structure plus biochemical reconstitution plus in vivo validation, defines molecular mechanism of multienzyme complex assembly","pmids":["39169182"],"is_preprint":false},{"year":2024,"finding":"METAP1 overexpression in human umbilical vein endothelial cells decreases tube formation (66%) and cell proliferation (72%), decreases VEGFA expression, and increases expression of preeclampsia-related genes (FLT1, INHBA, IL1B). METAP1 knockdown produces opposite effects, establishing antiangiogenic and proinflammatory roles in endothelial cells.","method":"Gain- and loss-of-function genetic approaches in HUVECs, tube formation assay, proliferation assay, gene expression analysis","journal":"Circulation research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — bidirectional loss/gain-of-function with multiple cellular readouts, single lab","pmids":["39727051"],"is_preprint":false},{"year":2025,"finding":"Human NMT1 exchanges with METAP1 at the ribosomal tunnel exit to form an active cotranslational complex together with NAC. NMT1 binding is sequence-selective and triggered by methionine excision by METAP1, which exposes the N-myristoylation motif in the nascent chain, enabling sequential cotranslational N-myristoylation.","method":"Biochemical interaction assays, cryo-EM structural studies, in vivo functional studies","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — cryo-EM structure plus biochemical reconstitution plus in vivo functional studies demonstrating exchange mechanism","pmids":["40639378"],"is_preprint":false},{"year":2025,"finding":"NAC recruits MetAP1 and NatD (NAA40) to ribosomes to form a multienzyme complex for sequential cotranslational modification of histones H2A and H4: MetAP1 excises the initiator methionine, then NatD acetylates the exposed N-terminus. MetAP1 and NatD cooperate in a confined ribosomal environment to enable efficient histone maturation.","method":"Cryo-EM structural studies, biochemical assays","journal":"Science advances","confidence":"High","confidence_rationale":"Tier 1 / Moderate — cryo-EM structure plus biochemical assays defining the multienzyme complex and sequential reaction mechanism","pmids":["41417911"],"is_preprint":false},{"year":2025,"finding":"Zinc activates MetAP1 via the metallochaperone ZNG1 (ZNG1-METAP1 complex), increasing intracellular SAM production. This promotes PRMT5-mediated symmetrical dimethylarginine (SDMA) methylation of AKT at R391 and R15, facilitating AKT translocation to the plasma membrane, interaction with mTORC2, and AKT activation to support cell proliferation and gut barrier function.","method":"Co-immunoprecipitation, SAM metabolite measurement, mass spectrometry for SDMA modification, AKT localization assay, cell proliferation assay","journal":"Advanced science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple biochemical and cell biology methods in single lab establishing pathway, but complexity of the pathway means each step has limited individual validation depth as reported in abstract","pmids":["40642900"],"is_preprint":false},{"year":2024,"finding":"Proteins whose N-termini are processed by MetAP1 (not MetAP2) are unaffected by MetAP2 loss or inhibition in the context of the UBR4-dependent Arg/N-degron pathway, demonstrating substrate specificity partitioning between MetAP1 and MetAP2 for N-degron pathway entry.","method":"Reporter assays, CRISPR-Cas9 knockout of MetAP2, bioinformatic analysis of endogenous substrates","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — reporter assays and CRISPR KO in single preprint, not yet peer-reviewed","pmids":["bio_10.1101_2024.10.03.616566"],"is_preprint":true}],"current_model":"METAP1 is a cotranslational metalloenzyme that removes the initiator methionine from newly synthesized proteins at the ribosomal tunnel exit; in eukaryotes, this activity is regulated by the nascent polypeptide-associated complex (NAC), which recruits METAP1 to the ribosome via a flexible tail, coordinates its action with downstream NatA and NatD acetyltransferases and NMT1 myristoyltransferase in substrate-specific multienzyme complexes, and relies on Zn2+ delivery by the GTP-dependent metallochaperone Zng1/ZNG1; the N-terminal zinc finger domain of METAP1 is required for 60S ribosome association and full catalytic efficiency, three cobalt ions are required for optimal catalysis of the human enzyme, and METAP1 function is essential for cell proliferation (particularly G2/M phase progression) and, together with MetAP2, for overall cellular viability."},"narrative":{"mechanistic_narrative":"METAP1 is a cobalt/zinc-dependent methionine aminopeptidase that cotranslationally removes the initiator methionine from nascent polypeptides emerging at the ribosomal tunnel exit, a processing step essential for protein maturation and cell viability [PMID:8618900, PMID:11968008]. In yeast, MAP1 deletion is viable but slow-growing while the map1/map2 double null is lethal, establishing that N-terminal methionine excision is essential and shared between two MetAPs that carry partly distinct in vivo substrate specificities — MetAP1 acting as the dominant isoform with characteristic preferences for small P1' residues and reduced activity toward Met-Val and Met-Thr substrates [PMID:8618900, PMID:11811952, PMID:20521764]. Human MetAP1 functionally substitutes for yeast Map1p, is required for cell proliferation and normal G2/M progression, and acts together with MetAP2 for overall viability [PMID:12144506, PMID:15962312, PMID:17114291]. Catalysis depends on an active-site geometry distinct from MetAP2: the human enzyme requires three Co2+ ions for optimal activity, with His212 coordinating a third metal unique to this enzyme, and its smaller active site underlies its resistance to the MetAP2 inhibitors fumagillin and ovalicin [PMID:9177176, PMID:16274222, PMID:17929833]; an N-terminal zinc-binding domain that unfolds upon zinc chelation mediates 60S ribosome association and full processing efficiency [PMID:11968008, PMID:25921012], with metal loading supplied by the GTP-dependent metallochaperone Zng1/ZNG1, which physically interacts with and transfers Zn2+/Co2+ to apo-enzyme [PMID:35584675]. Ribosomal recruitment is controlled by the nascent polypeptide-associated complex (NAC), which tethers METAP1 via a flexible tail and nucleates substrate-specific multienzyme complexes that coordinate methionine excision with downstream N-terminal acetylation by NatA and NatD and N-myristoylation by NMT1, the latter triggered by METAP1-dependent exposure of the myristoylation motif [PMID:37347872, PMID:39169182, PMID:40639378, PMID:41417911]. METAP1-processed N-termini also partition substrates into the Arg/N-degron pathway distinct from MetAP2 [PMID:bio_10.1101_2024.10.03.616566].","teleology":[{"year":1995,"claim":"Established that N-terminal methionine excision is an essential cellular function carried out cotranslationally by two redundant MetAPs, defining MetAP1 as a distinct eukaryotic enzyme bearing an N-terminal zinc finger absent in prokaryotes.","evidence":"Genetic deletion and complementation with immunoaffinity purification and enzymatic assay in S. cerevisiae","pmids":["8618900"],"confidence":"High","gaps":["Did not resolve the structural basis of substrate selection","Mechanism of ribosome targeting not addressed"]},{"year":1997,"claim":"Defined the catalytic and pharmacological distinction between the two isoforms — MetAP1 is not inhibited by fumagillin (a MetAP2-selective agent) and cannot substitute for blocked MetAP2 — and identified Asp219 as catalytically essential via a dominant-negative active-site mutation.","evidence":"In vivo yeast growth assays with fumagillin and site-directed mutagenesis with enzymatic assay","pmids":["9177176","9367524"],"confidence":"High","gaps":["Molecular basis of inhibitor selectivity not yet structurally explained","In vivo substrate spectrum unquantified"]},{"year":2002,"claim":"Demonstrated that the N-terminal zinc finger mediates 60S/80S ribosome association required for efficient processing, and that MetAP1 is the dominant in vivo methionine-excision isoform with isoform-specific cleavage preferences.","evidence":"Ribosome sedimentation profiling, zinc finger mutagenesis, and in vivo reporter processing assays in yeast deletion strains","pmids":["11968008","11811952"],"confidence":"High","gaps":["Did not identify the ribosomal docking partner","How the zinc finger contacts the 60S subunit unresolved"]},{"year":2002,"claim":"Confirmed functional conservation between yeast and human MetAP1, validating yeast as a model for the human enzyme.","evidence":"Heterologous complementation of yeast map1 null by human MetAP1","pmids":["12144506"],"confidence":"Medium","gaps":["Single complementation method without biochemical characterization of the human enzyme in this context"]},{"year":2003,"claim":"Connected MetAP1 to methionine salvage metabolism and pinned the molecular determinant of ovalicin selectivity to a single active-site residue (Thr in MetAP1 vs Ala in MetAP2).","evidence":"Genetic deletion strains with MET reporter and growth assays; bidirectional site-directed mutagenesis with in vivo inhibitor sensitivity testing","pmids":["12874831","14676204"],"confidence":"High","gaps":["Metabolic role characterized only in yeast","Structural consequences of the residue swap not yet visualized"]},{"year":2005,"claim":"Resolved the structural basis of MetAP1 inhibitor resistance — a smaller active site than MetAP2 — and identified Pro-x-x-Pro motifs consistent with ribosome binding.","evidence":"X-ray crystallography of human MetAP1 with comparative structural analysis","pmids":["16274222"],"confidence":"High","gaps":["Static structure does not capture nascent-chain engagement","Functional role of the Pro-x-x-Pro motifs not directly tested"]},{"year":2005,"claim":"Established that human MetAP1 is required for cell proliferation and that combined MetAP1/MetAP2 loss is near-lethal in human cells, recapitulating the yeast double-null phenotype.","evidence":"siRNA knockdown with proliferation assays in HUVEC and A549 cells","pmids":["15962312"],"confidence":"Medium","gaps":["Mechanism linking processing to proliferation not defined","Off-target effects of siRNA not fully excluded"]},{"year":2006,"claim":"Linked MetAP1 catalytic activity to G2/M cell cycle progression and demonstrated on-target action of selective inhibitors through rescue by MetAP1 overexpression and substrate-processing readouts.","evidence":"Selective inhibitors, FACS cell cycle analysis, siRNA, overexpression rescue, N-terminal processing assay, and ovalicin-MetAP1 co-crystal structure","pmids":["17114291","16823043"],"confidence":"High","gaps":["The specific substrate(s) driving the G2/M requirement not identified","How processing defects translate to cell cycle arrest unknown"]},{"year":2007,"claim":"Defined the unusual metal requirement of human MetAP1 — three Co2+ ions with His212 coordinating a unique third ion — distinguishing it from all other MetAP family members.","evidence":"Kinetic Co2+-activation analysis and active-site mutagenesis (H212A, H212K) with in vitro enzymatic assays","pmids":["17929833"],"confidence":"High","gaps":["Physiological identity of the in-cell catalytic metal not settled","How metals are loaded onto the enzyme in vivo not addressed here"]},{"year":2010,"claim":"Systematically defined human MetAP1 substrate specificity rules (small P1' residues, disfavoring Pro at P2' and acidic residues downstream), quantitatively separating its substrate space from MetAP2.","evidence":"Combinatorial peptide library screening with kinetic analysis of individual substrates","pmids":["20521764"],"confidence":"High","gaps":["Peptide specificity may not fully reflect cotranslational nascent-chain context","Endogenous substrate repertoire not enumerated"]},{"year":2015,"claim":"Provided structural characterization of the isolated N-terminal zinc-binding domain, confirming it folds in a zinc-dependent manner.","evidence":"NMR chemical shift assignment and EDTA perturbation of the isolated ZBD","pmids":["25921012"],"confidence":"Medium","gaps":["Full domain structure not solved","Functional ribosome-binding role not validated in this study"]},{"year":2016,"claim":"Mapped in vivo co-dependence of MetAP1 and MetAP2 on M[VT]X substrates at the proteome scale and linked fumagillin sensitivity to MetAP1 levels and glutathione redox state.","evidence":"N-terminomics and proteo-transcriptomic profiling across multiple cell lines with fumagillin treatment","pmids":["27542228"],"confidence":"Medium","gaps":["Mechanistic link between methionine excision and glutathione homeostasis not established","Single-lab proteomic dataset"]},{"year":2022,"claim":"Identified the GTP-dependent metallochaperone Zng1/ZNG1 as the in vivo source of metal for MetAP1, resolving how the apo-enzyme is loaded with Zn2+/Co2+.","evidence":"In vitro metal transfer and GTPase assays, yeast deletion genetics, and physical interaction/pulldown assays","pmids":["35584675"],"confidence":"High","gaps":["Selectivity between Zn2+ and Co2+ delivery in cells not resolved","Structural basis of the transfer reaction not determined"]},{"year":2023,"claim":"Defined how METAP1 accesses the ribosome — recruitment by NAC via a flexible tail to assemble an active methionine-excision complex at the tunnel exit while sparing ER-targeted proteins.","evidence":"Cryo-EM structural studies with biochemical interaction assays and in vivo validation","pmids":["37347872"],"confidence":"High","gaps":["How NAC discriminates cytosolic from ER substrates mechanistically not fully detailed","Stoichiometry and dynamics on actively translating ribosomes not resolved"]},{"year":2023,"claim":"Identified MetAP1 as a cisplatin-binding protein, implicating it in drug cytotoxicity through cysteine chelation.","evidence":"Competitive activity-based protein profiling with functional cytotoxicity validation","pmids":["37654507"],"confidence":"Medium","gaps":["Cysteine residue(s) targeted not pinpointed in the abstract","Contribution to cisplatin response relative to other targets unquantified"]},{"year":2024,"claim":"Showed that NAC orchestrates assembly of sequential cotranslational multienzyme complexes, pre-positioning MetAP1 with NatA (and releasing inhibitory HYPK) to couple methionine excision to N-terminal acetylation.","evidence":"Cryo-EM structures with biochemical reconstitution and in vivo studies","pmids":["39169182"],"confidence":"High","gaps":["How substrate sequence selects between alternative downstream enzymes not fully resolved","Kinetic ordering on native ribosomes inferred from reconstitution"]},{"year":2024,"claim":"Established cell-physiological roles of METAP1 in endothelium — antiangiogenic and proinflammatory effects modulating VEGFA and preeclampsia-related genes.","evidence":"Bidirectional gain- and loss-of-function in HUVECs with tube formation, proliferation, and gene expression assays","pmids":["39727051"],"confidence":"Medium","gaps":["Whether these effects require catalytic methionine excision unknown","Direct molecular targets linking METAP1 to VEGFA not identified"]},{"year":2024,"claim":"Demonstrated that MetAP1-specific N-terminal processing partitions substrates into the UBR4-dependent Arg/N-degron pathway independently of MetAP2.","evidence":"Reporter assays and CRISPR-Cas9 MetAP2 knockout with bioinformatic substrate analysis (preprint)","pmids":["bio_10.1101_2024.10.03.616566"],"confidence":"Medium","gaps":["Not yet peer-reviewed","Endogenous degron substrate set requires biochemical confirmation"]},{"year":2025,"claim":"Extended the cotranslational multienzyme paradigm to N-myristoylation and histone maturation — NMT1 exchanges with METAP1 after methionine excision exposes the myristoylation motif, and METAP1 cooperates with NatD for histone H2A/H4 N-terminal acetylation.","evidence":"Cryo-EM structures with biochemical reconstitution and in vivo functional studies","pmids":["40639378","41417911"],"confidence":"High","gaps":["Regulation of enzyme exchange order on individual nascent chains not resolved","Determinants selecting myristoylation vs acetylation routes not fully mapped"]},{"year":2025,"claim":"Connected ZNG1-dependent zinc activation of METAP1 to a signaling axis — increased SAM driving PRMT5-mediated AKT methylation, mTORC2 association, and AKT activation supporting proliferation and gut barrier function.","evidence":"Co-IP, SAM metabolite measurement, mass spectrometry of SDMA, AKT localization, and proliferation assays","pmids":["40642900"],"confidence":"Medium","gaps":["Mechanistic link from METAP1 activity to elevated SAM not directly established","Each pathway step has limited individual validation depth"]},{"year":null,"claim":"It remains unresolved which endogenous nascent-chain substrates drive the proliferation, G2/M, and signaling phenotypes of human METAP1, and how the NAC-organized multienzyme complexes are dynamically selected on individual translating ribosomes.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No causal substrate identified for the proliferation/G2/M requirement","Selection logic among NatA/NatD/NMT1 partner complexes on native polysomes unresolved","In-cell physiological catalytic metal identity not settled"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016787","term_label":"hydrolase activity","supporting_discovery_ids":[0,2,12,13]},{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,4,13]}],"localization":[{"term_id":"GO:0005840","term_label":"ribosome","supporting_discovery_ids":[3,17]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[17]}],"pathway":[{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[0,17,19]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[11]}],"complexes":["NAC-METAP1-NatA cotranslational complex","NAC-METAP1-NatD complex","NAC-METAP1-NMT1 complex","ZNG1-METAP1 complex"],"partners":["NACA","BTF3","NAA40","NMT1","ZNG1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P53582","full_name":"Methionine aminopeptidase 1","aliases":["Peptidase M 1"],"length_aa":386,"mass_kda":43.2,"function":"Cotranslationally removes the N-terminal methionine from nascent proteins. The N-terminal methionine is often cleaved when the second residue in the primary sequence is small and uncharged (Met-Ala-, Cys, Gly, Pro, Ser, Thr, or Val). Required for normal progression through the cell cycle","subcellular_location":"Cytoplasm","url":"https://www.uniprot.org/uniprotkb/P53582/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/METAP1","classification":"Not Classified","n_dependent_lines":744,"n_total_lines":1208,"dependency_fraction":0.6158940397350994},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"MAP1LC3B","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/METAP1","total_profiled":1310},"omim":[{"mim_id":"610151","title":"METHIONYL AMINOPEPTIDASE 1; METAP1","url":"https://www.omim.org/entry/610151"},{"mim_id":"601870","title":"METHIONINE AMINOPEPTIDASE 2; METAP2","url":"https://www.omim.org/entry/601870"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Cytosol","reliability":"Approved"},{"location":"Nucleoplasm","reliability":"Additional"},{"location":"Plasma membrane","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in 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Deletion of MAP1 is viable but causes slow growth; the double map1/map2 null is nonviable, establishing that N-terminal methionine removal is an essential function requiring two MetAPs.\",\n      \"method\": \"Genetic deletion, complementation assay, immunoaffinity purification, enzymatic activity assay\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — genetic epistasis (double null lethality), biochemical purification and enzymatic assay, replicated across multiple labs subsequently\",\n      \"pmids\": [\"8618900\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1997,\n      \"finding\": \"Fumagillin selectively inhibits S. cerevisiae MetAP2 in vivo but does NOT inhibit MetAP1 in vivo; MetAP1 function is insufficient to substitute for MetAP2 when MetAP2 is covalently blocked by fumagillin.\",\n      \"method\": \"In vivo yeast growth assay with fumagillin, differential inhibition of MetAP1 vs MetAP2\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo yeast genetic and pharmacological experiments, replicated across subsequent studies\",\n      \"pmids\": [\"9177176\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1997,\n      \"finding\": \"Mutation of the conserved cobalt-coordinating residue Asp219 (analogous to Asp97 in E. coli MetAP) in yeast MetAP1 to Asn reduces catalytic activity ~1000-fold and alters substrate specificity, demonstrating that Asp219 is essential for catalysis. The D219N mutant acts as a dominant negative, interfering with both wild-type MetAP1 and MetAP2 function in yeast.\",\n      \"method\": \"Site-directed mutagenesis, enzymatic activity assay, in vivo yeast growth assay\",\n      \"journal\": \"Archives of biochemistry and biophysics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — active-site mutagenesis combined with in vitro enzymatic assay and in vivo dominant-negative phenotype, single lab but multiple orthogonal methods\",\n      \"pmids\": [\"9367524\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Yeast MetAP1 is a ribosome-associated protein that primarily associates with the 60S ribosomal subunit and 80S translational complex. The N-terminal zinc finger domain is required for this association: single point mutations in the first or second zinc finger motif disrupt association with the 60S subunit and 80S complex and reduce N-terminal methionine removal from a reporter protein from ~100% to 31–35%.\",\n      \"method\": \"Ribosome sedimentation profiling, zinc finger mutagenesis, reporter protein N-terminal processing assay\",\n      \"journal\": \"Journal of cellular biochemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ribosome fractionation plus mutagenesis plus functional processing assay, multiple orthogonal methods in one study\",\n      \"pmids\": [\"11968008\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"MetAP1 plays the dominant role in N-terminal methionine removal in S. cerevisiae in vivo. Both MetAP1 and MetAP2 are less efficient when the penultimate residue is Val; MetAP2 is less efficient than MetAP1 for Gly, Cys, or Thr at this position, establishing different in vivo cleavage specificities.\",\n      \"method\": \"In vivo N-terminal processing assay of mutant GST reporter proteins in map1, map2, and wild-type yeast strains\",\n      \"journal\": \"Archives of biochemistry and biophysics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — systematic in vivo substrate specificity profiling with multiple deletion strains and multiple substrates\",\n      \"pmids\": [\"11811952\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Human MetAP1 functionally complements yeast map1 null growth defect, demonstrating conserved function between yeast and human MetAP1 in vivo.\",\n      \"method\": \"Heterologous expression of human MetAP1 in yeast map1 null strain, complementation growth assay\",\n      \"journal\": \"Protein and peptide letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — single lab, single complementation method\",\n      \"pmids\": [\"12144506\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"Yeast MetAP1 plays a significant role in methionine salvage metabolism, preventing premature activation of MET genes. In cells lacking MetAP1, excess methionine causes growth inhibition by product inhibition of MetAP2 (not MetAP1), revealing differential regulation and a functional distinction between the two isoforms.\",\n      \"method\": \"Genetic deletion strains, MET gene reporter assays, growth inhibition assays with methionine supplementation\",\n      \"journal\": \"Journal of cellular biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple genetic and metabolic assays in single lab\",\n      \"pmids\": [\"12874831\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"A single active-site residue difference between MetAP1 and MetAP2 accounts for differential sensitivity to ovalicin: the analogous position to MetAP2 Ala362 in MetAP1 is naturally a threonine, conferring resistance. Mutating MetAP1 Thr to Ala renders MetAP1 ovalicin-sensitive, and mutating MetAP2 Ala362 to Thr confers ovalicin resistance.\",\n      \"method\": \"Yeast-based mutagenesis screen, site-directed mutagenesis, in vivo inhibitor sensitivity assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — bidirectional mutagenesis (MetAP1 and MetAP2) with functional validation in yeast, defines molecular basis of specificity\",\n      \"pmids\": [\"14676204\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Crystal structure of human MetAP1 reveals that its active site is smaller than that of human MetAP2, explaining why ovalicin/fumagillin preferentially targets MetAP2. The N-terminal region of MetAP1 contains three Pro-x-x-Pro motifs consistent with ribosome binding.\",\n      \"method\": \"X-ray crystallography, structural comparison with MetAP2–ovalicin complex\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — crystal structure at defined resolution with comparative structural analysis explaining functional differences\",\n      \"pmids\": [\"16274222\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"siRNA-mediated knockdown of MetAP1 significantly inhibits proliferation of human umbilical vein endothelial cells (HUVEC, 70–80% inhibition) and A549 lung carcinoma cells (20–30%). Combined knockdown of MetAP1 and MetAP2 produces near-complete growth inhibition, mirroring the map1/map2 double null yeast phenotype.\",\n      \"method\": \"siRNA knockdown, cell proliferation assay\",\n      \"journal\": \"Journal of cellular biochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — siRNA knockdown with quantitative proliferation readout, multiple cell lines tested\",\n      \"pmids\": [\"15962312\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Ovalicin forms a low-affinity covalent adduct with the active-site histidine of human MetAP1 (His310), analogous to the covalent modification of His231 in MetAP2, but with different inhibitor alignment. Several active-site residues must shift outward to accommodate the inhibitor, explaining the lower affinity for MetAP1.\",\n      \"method\": \"X-ray crystallography at 1.1 Å resolution of ovalicin–MetAP1 complex\",\n      \"journal\": \"Protein science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — high-resolution crystal structure of inhibitor–enzyme complex with structural comparison\",\n      \"pmids\": [\"16823043\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Human MetAP1 (HsMetAP1) is required for normal G2/M phase cell cycle progression. Selective MetAP1 inhibitors (pyridine-2-carboxylic acid class) cause G2/M accumulation in tumor cells, induce apoptosis in leukemia lines, and cause N-terminal methionine retention in a known MetAP substrate. Overexpression of HsMetAP1 (but not MetAP2) confers resistance, and siRNA knockdown of HsMetAP1 recapitulates slow G2/M progression.\",\n      \"method\": \"Enzymatic assay, X-ray crystallography, cell cycle FACS analysis, siRNA knockdown, overexpression rescue, N-terminal processing assay\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — multiple orthogonal methods (structure, enzymatic assay, siRNA, overexpression rescue, cell cycle analysis) in single study\",\n      \"pmids\": [\"17114291\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Human cytosolic MetAP1 requires at least three Co2+ ions for optimal catalysis (Hill coefficient n≈2.9). The conserved residue His212 coordinates a third Co2+ ion unique to this enzyme; H212A and H212K mutations reduce kcat 60- and 1800-fold respectively, increase K0.5 for Co2+, and decrease cooperativity, establishing that three metal ions are functionally required—more than any other MetAP family member.\",\n      \"method\": \"Kinetic analysis, site-directed mutagenesis, Co2+-activation curves, in vitro enzymatic assay\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — rigorous kinetic analysis combined with active-site mutagenesis of two independent mutations, multiple kinetic parameters measured\",\n      \"pmids\": [\"17929833\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Systematic profiling of human MetAP1 substrate specificity using a combinatorial peptide library and kinetic analysis reveals: MetAP1 requires small residues (Gly, Ala, Ser, Cys, Pro, Thr, Val) at P1'; has lower activity toward Met-Val and Met-Thr substrates compared with MetAP2; is poorly active toward peptides with Pro at P2'; and disfavors acidic residues at P2'–P5'.\",\n      \"method\": \"Combinatorial peptide library screening, kinetic analysis of individual peptide substrates\",\n      \"journal\": \"Biochemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — systematic library screen plus individual kinetic measurements, defines substrate specificity rules\",\n      \"pmids\": [\"20521764\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"The N-terminal zinc-binding domain (ZBD, residues 1–83) of human MetAP1 contains two α-helical fragments (residues 44–49 and 59–82) and unfolds upon EDTA chelation of zinc, as demonstrated by NMR chemical shift assignments.\",\n      \"method\": \"NMR spectroscopy (15N-HSQC, chemical shift assignment), EDTA perturbation\",\n      \"journal\": \"Biomolecular NMR assignments\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — NMR structural characterization, single lab, provides partial structural information of isolated domain without full structure or functional validation\",\n      \"pmids\": [\"25921012\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Both MetAP1 and MetAP2 are required in vivo for N-terminal methionine excision from M[VT]X-class substrates. Cell sensitivity to fumagillin (MetAP2 inhibitor) correlates with MetAP1 protein levels and with the ability to modulate glutathione homeostasis; fumagillin-sensitive cells show glutathione redox alterations absent in resistant cells.\",\n      \"method\": \"Large-scale N-terminus proteomics (N-terminomics), fumagillin treatment in multiple cell lines, proteo-transcriptomic analysis\",\n      \"journal\": \"Oncotarget\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — global N-terminome profiling in multiple cell lines, single lab\",\n      \"pmids\": [\"27542228\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Zng1 (human ortholog conserved) is a GTP-dependent metallochaperone that transfers Zn2+ or Co2+ to apo-MetAP1 (Map1p) in vitro, requiring GTP hydrolysis for metal transfer—unlike known copper chaperones. Deletion of ZNG1 in yeast causes defective Map1p function; Zng1 physically interacts with Map1p.\",\n      \"method\": \"In vitro metal transfer assay, GTPase assay, yeast deletion genetics, pulldown interaction assay, proteomics\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — in vitro reconstitution of metal transfer, genetic epistasis in yeast, physical interaction demonstrated, multiple orthogonal methods\",\n      \"pmids\": [\"35584675\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"In eukaryotes, the nascent polypeptide-associated complex (NAC) controls ribosome binding of METAP1. NAC recruits METAP1 via a long flexible tail and provides a platform for formation of an active methionine excision complex at the ribosomal tunnel exit, ensuring efficient methionine excision from cytosolic proteins while sparing ER-targeted proteins.\",\n      \"method\": \"Biochemical interaction assays, structural studies (cryo-EM), in vivo functional studies\",\n      \"journal\": \"Science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — cryo-EM structure plus biochemical reconstitution plus in vivo validation, mechanistically defines how METAP1 accesses ribosome\",\n      \"pmids\": [\"37347872\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"MetAP1 is a novel cisplatin-binding protein: cisplatin binds to MetAP1 via cysteine chelation and contributes to cisplatin's cytotoxicity, as demonstrated by competitive activity-based protein profiling and functional validation.\",\n      \"method\": \"Competitive activity-based protein profiling (ABPP), functional cytotoxicity validation\",\n      \"journal\": \"RSC chemical biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — competitive ABPP with functional validation, single lab, limited mechanistic depth in abstract\",\n      \"pmids\": [\"37654507\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"NAC assembles a multienzyme complex with MetAP1 and NatA early during translation, pre-positioning the active sites of both enzymes for sequential cotranslational processing (methionine excision then N-terminal acetylation) of nascent proteins. NAC also releases inhibitory interactions from the NatA regulatory protein HYPK to activate NatA on the ribosome.\",\n      \"method\": \"Biochemical assays, cryo-EM structural studies, in vivo studies\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM structure plus biochemical reconstitution plus in vivo validation, defines molecular mechanism of multienzyme complex assembly\",\n      \"pmids\": [\"39169182\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"METAP1 overexpression in human umbilical vein endothelial cells decreases tube formation (66%) and cell proliferation (72%), decreases VEGFA expression, and increases expression of preeclampsia-related genes (FLT1, INHBA, IL1B). METAP1 knockdown produces opposite effects, establishing antiangiogenic and proinflammatory roles in endothelial cells.\",\n      \"method\": \"Gain- and loss-of-function genetic approaches in HUVECs, tube formation assay, proliferation assay, gene expression analysis\",\n      \"journal\": \"Circulation research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — bidirectional loss/gain-of-function with multiple cellular readouts, single lab\",\n      \"pmids\": [\"39727051\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Human NMT1 exchanges with METAP1 at the ribosomal tunnel exit to form an active cotranslational complex together with NAC. NMT1 binding is sequence-selective and triggered by methionine excision by METAP1, which exposes the N-myristoylation motif in the nascent chain, enabling sequential cotranslational N-myristoylation.\",\n      \"method\": \"Biochemical interaction assays, cryo-EM structural studies, in vivo functional studies\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — cryo-EM structure plus biochemical reconstitution plus in vivo functional studies demonstrating exchange mechanism\",\n      \"pmids\": [\"40639378\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"NAC recruits MetAP1 and NatD (NAA40) to ribosomes to form a multienzyme complex for sequential cotranslational modification of histones H2A and H4: MetAP1 excises the initiator methionine, then NatD acetylates the exposed N-terminus. MetAP1 and NatD cooperate in a confined ribosomal environment to enable efficient histone maturation.\",\n      \"method\": \"Cryo-EM structural studies, biochemical assays\",\n      \"journal\": \"Science advances\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — cryo-EM structure plus biochemical assays defining the multienzyme complex and sequential reaction mechanism\",\n      \"pmids\": [\"41417911\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Zinc activates MetAP1 via the metallochaperone ZNG1 (ZNG1-METAP1 complex), increasing intracellular SAM production. This promotes PRMT5-mediated symmetrical dimethylarginine (SDMA) methylation of AKT at R391 and R15, facilitating AKT translocation to the plasma membrane, interaction with mTORC2, and AKT activation to support cell proliferation and gut barrier function.\",\n      \"method\": \"Co-immunoprecipitation, SAM metabolite measurement, mass spectrometry for SDMA modification, AKT localization assay, cell proliferation assay\",\n      \"journal\": \"Advanced science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple biochemical and cell biology methods in single lab establishing pathway, but complexity of the pathway means each step has limited individual validation depth as reported in abstract\",\n      \"pmids\": [\"40642900\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Proteins whose N-termini are processed by MetAP1 (not MetAP2) are unaffected by MetAP2 loss or inhibition in the context of the UBR4-dependent Arg/N-degron pathway, demonstrating substrate specificity partitioning between MetAP1 and MetAP2 for N-degron pathway entry.\",\n      \"method\": \"Reporter assays, CRISPR-Cas9 knockout of MetAP2, bioinformatic analysis of endogenous substrates\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — reporter assays and CRISPR KO in single preprint, not yet peer-reviewed\",\n      \"pmids\": [\"bio_10.1101_2024.10.03.616566\"],\n      \"is_preprint\": true\n    }\n  ],\n  \"current_model\": \"METAP1 is a cotranslational metalloenzyme that removes the initiator methionine from newly synthesized proteins at the ribosomal tunnel exit; in eukaryotes, this activity is regulated by the nascent polypeptide-associated complex (NAC), which recruits METAP1 to the ribosome via a flexible tail, coordinates its action with downstream NatA and NatD acetyltransferases and NMT1 myristoyltransferase in substrate-specific multienzyme complexes, and relies on Zn2+ delivery by the GTP-dependent metallochaperone Zng1/ZNG1; the N-terminal zinc finger domain of METAP1 is required for 60S ribosome association and full catalytic efficiency, three cobalt ions are required for optimal catalysis of the human enzyme, and METAP1 function is essential for cell proliferation (particularly G2/M phase progression) and, together with MetAP2, for overall cellular viability.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"METAP1 is a cobalt/zinc-dependent methionine aminopeptidase that cotranslationally removes the initiator methionine from nascent polypeptides emerging at the ribosomal tunnel exit, a processing step essential for protein maturation and cell viability [#0, #3]. In yeast, MAP1 deletion is viable but slow-growing while the map1/map2 double null is lethal, establishing that N-terminal methionine excision is essential and shared between two MetAPs that carry partly distinct in vivo substrate specificities — MetAP1 acting as the dominant isoform with characteristic preferences for small P1' residues and reduced activity toward Met-Val and Met-Thr substrates [#0, #4, #13]. Human MetAP1 functionally substitutes for yeast Map1p, is required for cell proliferation and normal G2/M progression, and acts together with MetAP2 for overall viability [#5, #9, #11]. Catalysis depends on an active-site geometry distinct from MetAP2: the human enzyme requires three Co2+ ions for optimal activity, with His212 coordinating a third metal unique to this enzyme, and its smaller active site underlies its resistance to the MetAP2 inhibitors fumagillin and ovalicin [#1, #8, #12]; an N-terminal zinc-binding domain that unfolds upon zinc chelation mediates 60S ribosome association and full processing efficiency [#3, #14], with metal loading supplied by the GTP-dependent metallochaperone Zng1/ZNG1, which physically interacts with and transfers Zn2+/Co2+ to apo-enzyme [#16]. Ribosomal recruitment is controlled by the nascent polypeptide-associated complex (NAC), which tethers METAP1 via a flexible tail and nucleates substrate-specific multienzyme complexes that coordinate methionine excision with downstream N-terminal acetylation by NatA and NatD and N-myristoylation by NMT1, the latter triggered by METAP1-dependent exposure of the myristoylation motif [#17, #19, #21, #22]. METAP1-processed N-termini also partition substrates into the Arg/N-degron pathway distinct from MetAP2 [#24].\",\n  \"teleology\": [\n    {\n      \"year\": 1995,\n      \"claim\": \"Established that N-terminal methionine excision is an essential cellular function carried out cotranslationally by two redundant MetAPs, defining MetAP1 as a distinct eukaryotic enzyme bearing an N-terminal zinc finger absent in prokaryotes.\",\n      \"evidence\": \"Genetic deletion and complementation with immunoaffinity purification and enzymatic assay in S. cerevisiae\",\n      \"pmids\": [\"8618900\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not resolve the structural basis of substrate selection\", \"Mechanism of ribosome targeting not addressed\"]\n    },\n    {\n      \"year\": 1997,\n      \"claim\": \"Defined the catalytic and pharmacological distinction between the two isoforms — MetAP1 is not inhibited by fumagillin (a MetAP2-selective agent) and cannot substitute for blocked MetAP2 — and identified Asp219 as catalytically essential via a dominant-negative active-site mutation.\",\n      \"evidence\": \"In vivo yeast growth assays with fumagillin and site-directed mutagenesis with enzymatic assay\",\n      \"pmids\": [\"9177176\", \"9367524\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular basis of inhibitor selectivity not yet structurally explained\", \"In vivo substrate spectrum unquantified\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Demonstrated that the N-terminal zinc finger mediates 60S/80S ribosome association required for efficient processing, and that MetAP1 is the dominant in vivo methionine-excision isoform with isoform-specific cleavage preferences.\",\n      \"evidence\": \"Ribosome sedimentation profiling, zinc finger mutagenesis, and in vivo reporter processing assays in yeast deletion strains\",\n      \"pmids\": [\"11968008\", \"11811952\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not identify the ribosomal docking partner\", \"How the zinc finger contacts the 60S subunit unresolved\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Confirmed functional conservation between yeast and human MetAP1, validating yeast as a model for the human enzyme.\",\n      \"evidence\": \"Heterologous complementation of yeast map1 null by human MetAP1\",\n      \"pmids\": [\"12144506\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single complementation method without biochemical characterization of the human enzyme in this context\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Connected MetAP1 to methionine salvage metabolism and pinned the molecular determinant of ovalicin selectivity to a single active-site residue (Thr in MetAP1 vs Ala in MetAP2).\",\n      \"evidence\": \"Genetic deletion strains with MET reporter and growth assays; bidirectional site-directed mutagenesis with in vivo inhibitor sensitivity testing\",\n      \"pmids\": [\"12874831\", \"14676204\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Metabolic role characterized only in yeast\", \"Structural consequences of the residue swap not yet visualized\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Resolved the structural basis of MetAP1 inhibitor resistance — a smaller active site than MetAP2 — and identified Pro-x-x-Pro motifs consistent with ribosome binding.\",\n      \"evidence\": \"X-ray crystallography of human MetAP1 with comparative structural analysis\",\n      \"pmids\": [\"16274222\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Static structure does not capture nascent-chain engagement\", \"Functional role of the Pro-x-x-Pro motifs not directly tested\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Established that human MetAP1 is required for cell proliferation and that combined MetAP1/MetAP2 loss is near-lethal in human cells, recapitulating the yeast double-null phenotype.\",\n      \"evidence\": \"siRNA knockdown with proliferation assays in HUVEC and A549 cells\",\n      \"pmids\": [\"15962312\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism linking processing to proliferation not defined\", \"Off-target effects of siRNA not fully excluded\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Linked MetAP1 catalytic activity to G2/M cell cycle progression and demonstrated on-target action of selective inhibitors through rescue by MetAP1 overexpression and substrate-processing readouts.\",\n      \"evidence\": \"Selective inhibitors, FACS cell cycle analysis, siRNA, overexpression rescue, N-terminal processing assay, and ovalicin-MetAP1 co-crystal structure\",\n      \"pmids\": [\"17114291\", \"16823043\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"The specific substrate(s) driving the G2/M requirement not identified\", \"How processing defects translate to cell cycle arrest unknown\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Defined the unusual metal requirement of human MetAP1 — three Co2+ ions with His212 coordinating a unique third ion — distinguishing it from all other MetAP family members.\",\n      \"evidence\": \"Kinetic Co2+-activation analysis and active-site mutagenesis (H212A, H212K) with in vitro enzymatic assays\",\n      \"pmids\": [\"17929833\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological identity of the in-cell catalytic metal not settled\", \"How metals are loaded onto the enzyme in vivo not addressed here\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Systematically defined human MetAP1 substrate specificity rules (small P1' residues, disfavoring Pro at P2' and acidic residues downstream), quantitatively separating its substrate space from MetAP2.\",\n      \"evidence\": \"Combinatorial peptide library screening with kinetic analysis of individual substrates\",\n      \"pmids\": [\"20521764\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Peptide specificity may not fully reflect cotranslational nascent-chain context\", \"Endogenous substrate repertoire not enumerated\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Provided structural characterization of the isolated N-terminal zinc-binding domain, confirming it folds in a zinc-dependent manner.\",\n      \"evidence\": \"NMR chemical shift assignment and EDTA perturbation of the isolated ZBD\",\n      \"pmids\": [\"25921012\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Full domain structure not solved\", \"Functional ribosome-binding role not validated in this study\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Mapped in vivo co-dependence of MetAP1 and MetAP2 on M[VT]X substrates at the proteome scale and linked fumagillin sensitivity to MetAP1 levels and glutathione redox state.\",\n      \"evidence\": \"N-terminomics and proteo-transcriptomic profiling across multiple cell lines with fumagillin treatment\",\n      \"pmids\": [\"27542228\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanistic link between methionine excision and glutathione homeostasis not established\", \"Single-lab proteomic dataset\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Identified the GTP-dependent metallochaperone Zng1/ZNG1 as the in vivo source of metal for MetAP1, resolving how the apo-enzyme is loaded with Zn2+/Co2+.\",\n      \"evidence\": \"In vitro metal transfer and GTPase assays, yeast deletion genetics, and physical interaction/pulldown assays\",\n      \"pmids\": [\"35584675\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Selectivity between Zn2+ and Co2+ delivery in cells not resolved\", \"Structural basis of the transfer reaction not determined\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Defined how METAP1 accesses the ribosome — recruitment by NAC via a flexible tail to assemble an active methionine-excision complex at the tunnel exit while sparing ER-targeted proteins.\",\n      \"evidence\": \"Cryo-EM structural studies with biochemical interaction assays and in vivo validation\",\n      \"pmids\": [\"37347872\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How NAC discriminates cytosolic from ER substrates mechanistically not fully detailed\", \"Stoichiometry and dynamics on actively translating ribosomes not resolved\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Identified MetAP1 as a cisplatin-binding protein, implicating it in drug cytotoxicity through cysteine chelation.\",\n      \"evidence\": \"Competitive activity-based protein profiling with functional cytotoxicity validation\",\n      \"pmids\": [\"37654507\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Cysteine residue(s) targeted not pinpointed in the abstract\", \"Contribution to cisplatin response relative to other targets unquantified\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Showed that NAC orchestrates assembly of sequential cotranslational multienzyme complexes, pre-positioning MetAP1 with NatA (and releasing inhibitory HYPK) to couple methionine excision to N-terminal acetylation.\",\n      \"evidence\": \"Cryo-EM structures with biochemical reconstitution and in vivo studies\",\n      \"pmids\": [\"39169182\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How substrate sequence selects between alternative downstream enzymes not fully resolved\", \"Kinetic ordering on native ribosomes inferred from reconstitution\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Established cell-physiological roles of METAP1 in endothelium — antiangiogenic and proinflammatory effects modulating VEGFA and preeclampsia-related genes.\",\n      \"evidence\": \"Bidirectional gain- and loss-of-function in HUVECs with tube formation, proliferation, and gene expression assays\",\n      \"pmids\": [\"39727051\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether these effects require catalytic methionine excision unknown\", \"Direct molecular targets linking METAP1 to VEGFA not identified\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Demonstrated that MetAP1-specific N-terminal processing partitions substrates into the UBR4-dependent Arg/N-degron pathway independently of MetAP2.\",\n      \"evidence\": \"Reporter assays and CRISPR-Cas9 MetAP2 knockout with bioinformatic substrate analysis (preprint)\",\n      \"pmids\": [\"bio_10.1101_2024.10.03.616566\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Not yet peer-reviewed\", \"Endogenous degron substrate set requires biochemical confirmation\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Extended the cotranslational multienzyme paradigm to N-myristoylation and histone maturation — NMT1 exchanges with METAP1 after methionine excision exposes the myristoylation motif, and METAP1 cooperates with NatD for histone H2A/H4 N-terminal acetylation.\",\n      \"evidence\": \"Cryo-EM structures with biochemical reconstitution and in vivo functional studies\",\n      \"pmids\": [\"40639378\", \"41417911\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Regulation of enzyme exchange order on individual nascent chains not resolved\", \"Determinants selecting myristoylation vs acetylation routes not fully mapped\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Connected ZNG1-dependent zinc activation of METAP1 to a signaling axis — increased SAM driving PRMT5-mediated AKT methylation, mTORC2 association, and AKT activation supporting proliferation and gut barrier function.\",\n      \"evidence\": \"Co-IP, SAM metabolite measurement, mass spectrometry of SDMA, AKT localization, and proliferation assays\",\n      \"pmids\": [\"40642900\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanistic link from METAP1 activity to elevated SAM not directly established\", \"Each pathway step has limited individual validation depth\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"It remains unresolved which endogenous nascent-chain substrates drive the proliferation, G2/M, and signaling phenotypes of human METAP1, and how the NAC-organized multienzyme complexes are dynamically selected on individual translating ribosomes.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No causal substrate identified for the proliferation/G2/M requirement\", \"Selection logic among NatA/NatD/NMT1 partner complexes on native polysomes unresolved\", \"In-cell physiological catalytic metal identity not settled\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016787\", \"supporting_discovery_ids\": [0, 2, 12, 13]},\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 4, 13]},\n      {\"term_id\": \"GO:0008270\", \"supporting_discovery_ids\": [3, 14]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005840\", \"supporting_discovery_ids\": [3, 17]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [17]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [0, 17, 19]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [11]}\n    ],\n    \"complexes\": [\"NAC-METAP1-NatA cotranslational complex\", \"NAC-METAP1-NatD complex\", \"NAC-METAP1-NMT1 complex\", \"ZNG1-METAP1 complex\"],\n    \"partners\": [\"NACA\", \"BTF3\", \"NAA40\", \"NMT1\", \"ZNG1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}