Biology:METAP2

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Short description: Protein-coding gene in humans


A representation of the 3D structure of the protein myoglobin showing turquoise α-helices.
Generic protein structure example

Methionine aminopeptidase 2 is an enzyme that in humans is encoded by the METAP2 gene.[1][2]

Methionine aminopeptidase 2, a member of the dimetallohydrolase family, is a cytosolic metalloenzyme that catalyzes the hydrolytic removal of N-terminal methionine residues from nascent proteins.[3][4][5]

  • peptide-methionine [math]\displaystyle{ \rightleftharpoons }[/math] peptide + methionine

MetAP2 is found in all organisms and is especially important because of its critical role in tissue repair and protein degradation.[3] Furthermore, MetAP2 is of particular interest because the enzyme plays a key role in angiogenesis, the growth of new blood vessels, which is necessary for the progression of diseases including solid tumor cancers and rheumatoid arthritis.[6] MetAP2 is also the target of two groups of anti-angiogenic natural products, ovalicin and fumagillin, and their analogs such as beloranib.[7][8][9][10]

Structure

In living organisms, the start codon that initiates protein synthesis codes for either methionine (eukaryotes) or formylmethionine (prokaryotes). In E. coli (prokaryote), an enzyme called formylmethionine deformylase can cleave the formyl group, leaving just the N-terminal methionine residue. For proteins with small, uncharged penultimate N-terminal residues, a methionine aminopeptidase can cleave the methionine residue.[3] The number of genes encoding for a methionine aminopeptidase varies between organisms. In E. coli, there is only one known MetAP, a 29,333 Da monomeric enzyme coded for by a gene consisting of 264 codons.[3] The knockout of this gene in E. coli leads to cell inviability.[11] In humans, there are two genes encoding MetAP, MetAP1 and MetAP2. MetAP1 codes for a 42 kDa enzyme, while MetAP2 codes for a 67 kDa enzyme. Yeast MetAP1 is 40 percent homologous to E. coli MetAP; within S. cerevisiae, MetAP2 is 22 percent homologous with the sequence of MetAP1; MetAP2 is highly conserved between S. cerevisiae and humans.[12] In contrast to prokaryotes, eukaryotic S. cerevisiae strains lacking the gene for either MetAP1 or MetAP2 are viable, but exhibit a slower growth rate than a control strain expressing both genes.

File:MetAP2 active site.ogv

Active site

The active site of MetAP2 has a structural motif characteristic of many metalloenzymes—including the dioxygen carrier protein, hemerythrin; the dinuclear non-heme iron protein, ribonucleotide reductase; leucine aminopeptidase; urease; arginase; several phosphatases and phosphoesterases—that includes two bridging carboxylate ligands and a bridging water or hydroxide ligand.[3][4][13][14][15][16][17] Specifically in human MetAP2 (PDB: 1BOA), one of the catalytic metal ions is bound to His331, Glu364, Glu459, Asp263, and a bridging water or hydroxide, while the other metal ion is bound to Asp251 (bidentate), App262 (bidentate), Glu459, and the same bridging water or hydroxide. Here, the two bridging carboxylates are Asp262 and Glu459.

Dimetal center

The identity of the active site metal ions under physiological conditions has not been successfully established, and remains a controversial issue. MetAP2 shows activity in the presence of Zn(II), Co(II), Mn(II), and Fe(II) ions, and various authors have argued any given metal ion is the physiological one: some in the presence of iron,[18] others in cobalt,[19][20] others in manganese,[21] and yet others in the presence of zinc.[22] Nonetheless, the majority of crystallographers have crystallized MetAP2 either in the presence of Zn(II) or Co(II) (see PDB database).

Mechanism

Figure 2. Two proposed reaction mechanisms for MetAP in E. coli. (A) Tetrahedral intermediate stabilized by Glu204 and metal center. (B) Tetrahedral intermediate stabilized His178 and metal center.[23]

The bridging water or hydroxide ligand acts as a nucleophile during the hydrolysis reaction, but the exact mechanism of catalysis is not yet known.[6][15][24] The catalytic mechanisms of hydrolase enzymes depend greatly on the identity of the bridging ligand,[25] which can be challenging to determine due to the difficulty of studying hydrogen atoms via x-ray crystallography.

The histidine residues shown in the mechanism to the right, H178 and H79, are conserved in all MetAPs (MetAP1s and MetAP2s) sequenced to date, suggesting their presence is important to catalytic activity.[26] Based upon X-ray crystallographic data, histidine 79 (H79) has been proposed to help position the methionine residue in the active site and transfer a proton to the newly exposed N-terminal amine.[8] Lowther and Colleagues have proposed two possible mechanisms for MetAP2 in E. coli, shown at the right.[10]

Function

While previous studies have indicated MetAP2 catalyzes the removal of N-terminal methionine residues in vitro, the function of this enzyme in vivo may be more complex. For example, a significant correlation exists between the inhibition of the enzymatic activity of MetAP2 and inhibition of cell growth, thus implicating the enzyme in endothelial cell proliferation.[9] For this reason, cancer researchers have singled out MetAP2 as a potential target for the inhibition of angiogenesis. Moreover, studies have demonstrated that MetAP2 copurifies and interacts with the α subunit of eukaryotic initiation factor 2 (eIF2), a protein that is necessary for protein synthesis in vivo.[27] Specifically, MetAP2 protects eIF-2α from inhibitory phosphorylation from the enzyme eIF-2α kinase, inhibits RNA-dependent protein kinase (PKR)-catalyzed eIF-2 R-subunit phosphorylation, and also reverses PKR-mediated inhibition of protein synthesis in intact cells.

Clinical significance

Figure 3. Fumagillin (green and red) bound to human MetAP2 active site (multicolored, with cyan, purple, and pink corresponding to helices, sheets, and loops, respectively), with dimetal ions (blue) shown.

Numerous studies implicate MetAP2 in angiogenesis.[9][16][28][29][30] Specifically, the covalent binding of either the ovalicin or fumagillin epoxide moiety to the active site histidine residue of MetAP2 has been shown to inactivate the enzyme, thereby inhibiting angiogenesis. The way in which MetAP2 regulates angiogenesis has yet to be established, however, such that further study is required to validate that antiangiogenic activity results directly from MetAP2 inhibition. Nevertheless, with both the growth and metastasis of solid tumors depending heavily on angiogenesis, fumagillin and its analogs—including TNP-470, caplostatin, and beloranib—as well as ovalicin represent potential anticancer agents.[29][30] Moreover, the ability of MetAP2 to decrease cell viability in prokaryotic and small eukaryotic organisms has made it a target for antibacterial agents.[9] Thus far, both fumagillin and TNP-470 have been shown to possess antimalarial activity both in vitro and in vivo, and fumarranol, another fumagillin analog, represents a promising lead.[30]

The fumagillin-derived METAP2 inhibitor beloranib (ZGN-433, CDK-732) has shown efficacy in reducing weight in severely obese subjects.[31] MetAP2 inhibitors work by re-establishing insulin sensitivity and balance to the ways the body metabolizes fat, leading to substantial loss of body weight. Development of beloranib was halted in 2016 after two deaths during clinical trials for patients with Praeder-Willi Syndrome.[32] A polymer-drug conjugate of a novel MetAP2 inhibitor called evexomostat being developed by SynDevRx, Inc. entered clinical development for late-stage cancer patients in 2016. Phase 1 dose escalation studies were completed in 2020. In 2022, SynDevRx initiated a Phase 2 clinical study of evexomostat in collaboration with Memorial Sloan Kettering Cancer Center of New York to assess the safety and efficacy in recurrent, metastatic triple-negative breast cancer in combination with the drug eribulin (Halaven(R)). In 2023, SynDevRx initiated another Phase 2 clinical study of evexomostat in combination with the alpelisib (Piqray (R)) and fulvestrant (Faslodex (R)) in metastatic HR+/Her2- breast cancer patients.

Interactions

METAP2 has been shown to interact with Protein kinase R.[33]

References

  1. ↑ "Eukaryotic methionyl aminopeptidases: two classes of cobalt-dependent enzymes". Proc Natl Acad Sci U S A 92 (17): 7714–8. September 1995. doi:10.1073/pnas.92.17.7714. PMID 7644482. Bibcode: 1995PNAS...92.7714A. 
  2. ↑ "Evidence that the human homologue of a rat initiation factor-2 associated protein (p67) is a methionine aminopeptidase". Biochem Biophys Res Commun 227 (1): 152–9. November 1996. doi:10.1006/bbrc.1996.1482. PMID 8858118. 
  3. ↑ 3.0 3.1 3.2 3.3 3.4 "EPR Studies on the Mono- and Dicobalt(II)-Substituted Forms of the Aminopeptidase from Aeromonas proteolytica. Insight into the Catalytic Mechanism of Dinuclear Hydrolases". J. Am. Chem. Soc. 119 (8): 1923–1933. 1997. doi:10.1021/ja963021v. https://epublications.marquette.edu/physics_fac/62. 
  4. ↑ 4.0 4.1 "Dicobalt II-II, II-III, and III-III complexes as spectroscopic models for dicobalt enzyme active sites". Inorg Chem 47 (12): 5079–92. June 2008. doi:10.1021/ic7020534. PMID 18494467. 
  5. ↑ "Magnetic circular dichroism and cobalt(II) binding equilibrium studies of Escherichia coli methionyl aminopeptidase". J. Am. Chem. Soc. 126 (39): 12316–24. October 2004. doi:10.1021/ja0485006. PMID 15453765. 
  6. ↑ 6.0 6.1 Folkman J (January 1995). "Angiogenesis in cancer, vascular, rheumatoid and other disease". Nat. Med. 1 (1): 27–31. doi:10.1038/nm0195-27. PMID 7584949. 
  7. ↑ Taunton J (July 1997). "How to starve a tumor". Chem. Biol. 4 (7): 493–6. doi:10.1016/S1074-5521(97)90320-3. PMID 9263636. 
  8. ↑ 8.0 8.1 "The anti-angiogenic agent fumagillin covalently binds and inhibits the methionine aminopeptidase, MetAP-2". Proc. Natl. Acad. Sci. U.S.A. 94 (12): 6099–103. June 1997. doi:10.1073/pnas.94.12.6099. PMID 9177176. Bibcode: 1997PNAS...94.6099S. 
  9. ↑ 9.0 9.1 9.2 9.3 "Methionine aminopeptidase (type 2) is the common target for angiogenesis inhibitors AGM-1470 and ovalicin". Chem. Biol. 4 (6): 461–71. June 1997. doi:10.1016/S1074-5521(97)90198-8. PMID 9224570. 
  10. ↑ 10.0 10.1 "The anti-angiogenic agent fumagillin covalently modifies a conserved active-site histidine in the Escherichia coli methionine aminopeptidase". Proc. Natl. Acad. Sci. U.S.A. 95 (21): 12153–7. October 1998. doi:10.1073/pnas.95.21.12153. PMID 9770455. Bibcode: 1998PNAS...9512153L. 
  11. ↑ "Methionine aminopeptidase gene of Escherichia coli is essential for cell growth". J. Bacteriol. 171 (7): 4071–2. July 1989. doi:10.1128/jb.171.7.4071-4072.1989. PMID 2544569. 
  12. ↑ "Amino-terminal protein processing in Saccharomyces cerevisiae is an essential function that requires two distinct methionine aminopeptidases". Proc. Natl. Acad. Sci. U.S.A. 92 (26): 12357–61. December 1995. doi:10.1073/pnas.92.26.12357. PMID 8618900. Bibcode: 1995PNAS...9212357L. 
  13. ↑ "Synthesis and spectroscopic studies of non-heme diiron(III) species with a terminal hydroperoxide ligand: models for hemerythrin". Inorg Chem 40 (18): 4662–73. August 2001. doi:10.1021/ic010076b. PMID 11511213. 
  14. ↑ "Supramolecular Control of Stepwise and Selective Carboxylate Ligand Substitution in Aqua-Carboxylato-Bridged Dimetal(II) Complexes". J. Am. Chem. Soc. 115 (26): 12617–12618. 1993. doi:10.1021/ja00079a064. 
  15. ↑ 15.0 15.1 "Magnetic, spectroscopic, and structural studies of dicobalt hydroxamates and model hydrolases". Inorg Chem 40 (23): 5962–71. November 2001. doi:10.1021/ic0103345. PMID 11681912. 
  16. ↑ 16.0 16.1 "Magnetic circular dichroism study of a dicobalt(II) methionine aminopeptidase/fumagillin complex and dicobalt II-II and II-III model complexes". Inorg Chem 47 (22): 10499–508. November 2008. doi:10.1021/ic8011553. PMID 18921993. 
  17. ↑ Wilcox DE (November 1996). "Binuclear Metallohydrolases". Chem. Rev. 96 (7): 2435–2458. doi:10.1021/cr950043b. PMID 11848832. 
  18. ↑ "The methionyl aminopeptidase from Escherichia coli can function as an iron(II) enzyme". Biochemistry 38 (34): 11079–85. August 1999. doi:10.1021/bi990872h. PMID 10460163. https://epublications.marquette.edu/cgi/viewcontent.cgi?article=1303&context=chem_fac. 
  19. ↑ "Molecular cloning, sequencing, deletion, and overexpression of a methionine aminopeptidase gene from Saccharomyces cerevisiae". J. Biol. Chem. 267 (12): 8007–11. April 1992. doi:10.1016/S0021-9258(18)42400-3. PMID 1569059. 
  20. ↑ "Characterization of native and recombinant forms of an unusual cobalt-dependent proline dipeptidase (prolidase) from the hyperthermophilic archaeon Pyrococcus furiosus". J. Bacteriol. 180 (18): 4781–9. September 1998. doi:10.1128/JB.180.18.4781-4789.1998. PMID 9733678. 
  21. ↑ "Physiologically relevant metal cofactor for methionine aminopeptidase-2 is manganese". Biochemistry 42 (17): 5035–42. May 2003. doi:10.1021/bi020670c. PMID 12718546. 
  22. ↑ "Which one among Zn(II), Co(II), Mn(II), and Fe(II) is the most efficient ion for the methionine aminopeptidase catalyzed reaction?". J. Am. Chem. Soc. 129 (25): 7776–84. June 2007. doi:10.1021/ja068168t. PMID 17523636. 
  23. ↑ "Insights into the mechanism of Escherichia coli methionine aminopeptidase from the structural analysis of reaction products and phosphorus-based transition-state analogues". Biochemistry 38 (45): 14810–9. November 1999. doi:10.1021/bi991711g. PMID 10555963. 
  24. ↑ "Electronic Paramagnetic Resonance and Magnetic Properties of Model Complexes for Binuclear Active Sites in Hydrolase Enzymes". Inorg. Chem. 36 (12): 2617–2622. 1997. doi:10.1021/ic960988r. 
  25. ↑ "Diiron(II) mu-aqua-mu-hydroxo model for non-heme iron sites in proteins". Inorg Chem 44 (24): 8656–8. November 2005. doi:10.1021/ic051739i. PMID 16296818. 
  26. ↑ "Mutations at the S1 sites of methionine aminopeptidases from Escherichia coli and Homo sapiens reveal the residues critical for substrate specificity". J. Biol. Chem. 279 (20): 21128–34. May 2004. doi:10.1074/jbc.M401679200. PMID 14976199. 
  27. ↑ "A eukaryotic translation initiation factor 2-associated 67 kDa glycoprotein partially reverses protein synthesis inhibition by activated double-stranded RNA-dependent protein kinase in intact cells". Biochemistry 35 (25): 8275–80. June 1996. doi:10.1021/bi953028+. PMID 8679583. 
  28. ↑ "An orally delivered small-molecule formulation with antiangiogenic and anticancer activity". Nat. Biotechnol. 26 (7): 799–807. July 2008. doi:10.1038/nbt1415. PMID 18587385. 
  29. ↑ 29.0 29.1 Sato Y (2004). "Aminopeptidases in Health and Disease: Role of Aminopeptidase in Angiogenesis". Biol. Pharm. Bull. 27 (6): 772–776. doi:10.1248/bpb.27.772. PMID 15187415. 
  30. ↑ 30.0 30.1 30.2 "Fumagillin and fumarranol interact with P. falciparum methionine aminopeptidase 2 and inhibit malaria parasite growth in vitro and in vivo". Chem. Biol. 16 (2): 193–202. February 2009. doi:10.1016/j.chembiol.2009.01.006. PMID 19246010. 
  31. ↑ "Zafgen Announces Positive Topline Phase 1b Data for ZGN-433 in Obesity". MedNews. Drugs.com. 2011-01-01. https://www.drugs.com/clinical_trials/zafgen-announces-positive-topline-phase-1b-data-zgn-433-obesity-10955.html. 
  32. ↑ "Zafgen Halts Development of Beloranib, to Cut Jobs by ~34%". nasdaq.com. July 20, 2016. http://www.nasdaq.com/article/zafgen-halts-development-of-beloranib-to-cut-jobs-by-34-cm651992. 
  33. ↑ "In vivo regulation of the dsRNA-dependent protein kinase PKR by the cellular glycoprotein p67". Biochemistry 39 (51): 16016–25. December 2000. doi:10.1021/bi001754t. PMID 11123929. 

Further reading



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