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Factor-inhibiting hypoxia-inducible factor (FIH) catalyses the post-translational hydroxylation of histidinyl residues within ankyrin repeat domains.

Yang M, Chowdhury R, Ge W, Hamed RB, McDonough MA, Claridge TD, Kessler BM, Cockman ME, Ratcliffe PJ, Schofield CJ - FEBS J. (2011)

Bottom Line: However, there are few reports on the selectivity of FIH for the hydroxylation of specific residues.NMR and crystallographic analyses show that the histidinyl hydroxylation occurs at the β-position.The results further expand the scope of FIH-catalysed hydroxylations.

View Article: PubMed Central - PubMed

Affiliation: Oxford Centre for Integrative Systems Biology, University of Oxford, Oxford, UK.

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Related in: MedlinePlus

Structure of the FIH complexes. (A) Surface representation of the FIH·TNKS2 538–558 dimer structure (PDB ID: 2Y0I) to 2.28 Å resolution showing apparent electron density for residues Ser 540 to His 553 of the TNKS2 538–558 peptide (2Fo − Fc map, contoured to 1σ). (B) Stereoview stick representation of the FIH active site of the FIH·TNKS2 538–558 complex (FIH, deep teal; TNKS2 538–558, yellow; Fe(II), orange). (C) Stereoview stick representation of the superimposed FIH·mNotch1 1930–1949 (PDB ID: 3P3N, FIH in green and N1 1930–1949 in white) and FIH·HIF-1αCAD 788–826 (PDB ID: 1H2K; FIH in purple and HIF-1αCAD 788-826 in salmon) complexes. A comparison of all FIH complexes suggests that the pro-S hydrogen of His 553 in TNKS2 538–558 is likely analogously positioned as that observed for hydroxylated asparagines in FIH·mNotch1 1930–1949 (PDB ID: 3P3N) and FIH·HIF-1αCAD 788–826 (PDB ID: 1H2K) complexes. (B) and (C) also illustrate the differences in side-chain conformation for Gln 239FIH and Tyr 103FIH between the FIH·TNKS2 538–558 and FIH·mNotch1 1930–1949/FIH·HIF-1αCAD 788-826 complexes.
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fig05: Structure of the FIH complexes. (A) Surface representation of the FIH·TNKS2 538–558 dimer structure (PDB ID: 2Y0I) to 2.28 Å resolution showing apparent electron density for residues Ser 540 to His 553 of the TNKS2 538–558 peptide (2Fo − Fc map, contoured to 1σ). (B) Stereoview stick representation of the FIH active site of the FIH·TNKS2 538–558 complex (FIH, deep teal; TNKS2 538–558, yellow; Fe(II), orange). (C) Stereoview stick representation of the superimposed FIH·mNotch1 1930–1949 (PDB ID: 3P3N, FIH in green and N1 1930–1949 in white) and FIH·HIF-1αCAD 788–826 (PDB ID: 1H2K; FIH in purple and HIF-1αCAD 788-826 in salmon) complexes. A comparison of all FIH complexes suggests that the pro-S hydrogen of His 553 in TNKS2 538–558 is likely analogously positioned as that observed for hydroxylated asparagines in FIH·mNotch1 1930–1949 (PDB ID: 3P3N) and FIH·HIF-1αCAD 788–826 (PDB ID: 1H2K) complexes. (B) and (C) also illustrate the differences in side-chain conformation for Gln 239FIH and Tyr 103FIH between the FIH·TNKS2 538–558 and FIH·mNotch1 1930–1949/FIH·HIF-1αCAD 788-826 complexes.

Mentions: To investigate how a His residue is hydroxylated by FIH, we crystallized FIH in complex with Fe(II), 2OG and the TNKS2 538–558 fragment under anaerobic conditions [19]. The resultant overall FIH structure (2.28 Å resolution, Fig. 5A) was similar to reported FIH structures (rmsd values of 0.30–0.33 Å for Cα backbone atoms) and reveals that the backbone of TNKS2 538–558 is bound to FIH in a manner that is similar overall to analogous ARD fragments that undergo Asn hydroxylation and a fragment of the HIF-1α CAD substrate (rmsd for Cα backbone atoms of TNKS2 versus mNotch-1 and CAD ∼ 0.2 Å) [6,19]. At the N-terminus of the bound TNKS2 538–558 fragment, residues 541–546 form a short α-helix, possibly reflecting that in the ankyrin repeat (AR) of the parent tankyrase ARD protein (Fig. 5A). At the active site, the Fe(II) and 2OG are bound as first reported for the structure of FIH in complex with Fe(II) and a fragment of the HIF-1αCAD [20]. In the FIH·TNKS2 538–558 structure, the β-methylene of His 553 is positioned such that the pro-S hydrogen of its methylene group projects towards the Fe(II) centre (Fig. 5B), suggesting that it is hydroxylated to give the 3S-hydroxy product, as observed for Asn hydroxylation by FIH [16]. Histidine binding at the FIH active site apparently induces a stacking interaction between the substrate imidazole and the phenolic rings of Tyr 102FIH and His 199FIH, which is one of the iron-complexing residues (Fig. 5B). Close to the β-methylene of His 553, we observed an electron density that was refined as a water molecule, although we cannot rule out the possibility that this density represents another species (e.g. partial reaction of the substrate; however, attempted refinements with hydroxylated His 553 were unsuccessful). The imidazole side chain of His 553 in the TNKS2 538–558 fragment points towards the γ-methylene of the side chain of Gln 239FIH (Fig. 5B). Previous structures have shown that Gln 239FIH binds via hydrogen-bonding interactions to the side chain of Asn residues that undergo hydroxylation, for example, Asn 803 of HIF-1α (Fig. 5C) [6,19]. However, in the TNKS2 538–558 structure, the side-chain amide of Gln 239FIH is moved away from the side chain of the hydroxylated residue (i.e. with His 553 compared with a hydroxylated Asn residue) such that it is positioned to make a hydrogen bond with the backbone amide of Tyr 102FIH. Apparently concomitant with this change, the side chain of Tyr 103FIH also moves (Fig. 5B).


Factor-inhibiting hypoxia-inducible factor (FIH) catalyses the post-translational hydroxylation of histidinyl residues within ankyrin repeat domains.

Yang M, Chowdhury R, Ge W, Hamed RB, McDonough MA, Claridge TD, Kessler BM, Cockman ME, Ratcliffe PJ, Schofield CJ - FEBS J. (2011)

Structure of the FIH complexes. (A) Surface representation of the FIH·TNKS2 538–558 dimer structure (PDB ID: 2Y0I) to 2.28 Å resolution showing apparent electron density for residues Ser 540 to His 553 of the TNKS2 538–558 peptide (2Fo − Fc map, contoured to 1σ). (B) Stereoview stick representation of the FIH active site of the FIH·TNKS2 538–558 complex (FIH, deep teal; TNKS2 538–558, yellow; Fe(II), orange). (C) Stereoview stick representation of the superimposed FIH·mNotch1 1930–1949 (PDB ID: 3P3N, FIH in green and N1 1930–1949 in white) and FIH·HIF-1αCAD 788–826 (PDB ID: 1H2K; FIH in purple and HIF-1αCAD 788-826 in salmon) complexes. A comparison of all FIH complexes suggests that the pro-S hydrogen of His 553 in TNKS2 538–558 is likely analogously positioned as that observed for hydroxylated asparagines in FIH·mNotch1 1930–1949 (PDB ID: 3P3N) and FIH·HIF-1αCAD 788–826 (PDB ID: 1H2K) complexes. (B) and (C) also illustrate the differences in side-chain conformation for Gln 239FIH and Tyr 103FIH between the FIH·TNKS2 538–558 and FIH·mNotch1 1930–1949/FIH·HIF-1αCAD 788-826 complexes.
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fig05: Structure of the FIH complexes. (A) Surface representation of the FIH·TNKS2 538–558 dimer structure (PDB ID: 2Y0I) to 2.28 Å resolution showing apparent electron density for residues Ser 540 to His 553 of the TNKS2 538–558 peptide (2Fo − Fc map, contoured to 1σ). (B) Stereoview stick representation of the FIH active site of the FIH·TNKS2 538–558 complex (FIH, deep teal; TNKS2 538–558, yellow; Fe(II), orange). (C) Stereoview stick representation of the superimposed FIH·mNotch1 1930–1949 (PDB ID: 3P3N, FIH in green and N1 1930–1949 in white) and FIH·HIF-1αCAD 788–826 (PDB ID: 1H2K; FIH in purple and HIF-1αCAD 788-826 in salmon) complexes. A comparison of all FIH complexes suggests that the pro-S hydrogen of His 553 in TNKS2 538–558 is likely analogously positioned as that observed for hydroxylated asparagines in FIH·mNotch1 1930–1949 (PDB ID: 3P3N) and FIH·HIF-1αCAD 788–826 (PDB ID: 1H2K) complexes. (B) and (C) also illustrate the differences in side-chain conformation for Gln 239FIH and Tyr 103FIH between the FIH·TNKS2 538–558 and FIH·mNotch1 1930–1949/FIH·HIF-1αCAD 788-826 complexes.
Mentions: To investigate how a His residue is hydroxylated by FIH, we crystallized FIH in complex with Fe(II), 2OG and the TNKS2 538–558 fragment under anaerobic conditions [19]. The resultant overall FIH structure (2.28 Å resolution, Fig. 5A) was similar to reported FIH structures (rmsd values of 0.30–0.33 Å for Cα backbone atoms) and reveals that the backbone of TNKS2 538–558 is bound to FIH in a manner that is similar overall to analogous ARD fragments that undergo Asn hydroxylation and a fragment of the HIF-1α CAD substrate (rmsd for Cα backbone atoms of TNKS2 versus mNotch-1 and CAD ∼ 0.2 Å) [6,19]. At the N-terminus of the bound TNKS2 538–558 fragment, residues 541–546 form a short α-helix, possibly reflecting that in the ankyrin repeat (AR) of the parent tankyrase ARD protein (Fig. 5A). At the active site, the Fe(II) and 2OG are bound as first reported for the structure of FIH in complex with Fe(II) and a fragment of the HIF-1αCAD [20]. In the FIH·TNKS2 538–558 structure, the β-methylene of His 553 is positioned such that the pro-S hydrogen of its methylene group projects towards the Fe(II) centre (Fig. 5B), suggesting that it is hydroxylated to give the 3S-hydroxy product, as observed for Asn hydroxylation by FIH [16]. Histidine binding at the FIH active site apparently induces a stacking interaction between the substrate imidazole and the phenolic rings of Tyr 102FIH and His 199FIH, which is one of the iron-complexing residues (Fig. 5B). Close to the β-methylene of His 553, we observed an electron density that was refined as a water molecule, although we cannot rule out the possibility that this density represents another species (e.g. partial reaction of the substrate; however, attempted refinements with hydroxylated His 553 were unsuccessful). The imidazole side chain of His 553 in the TNKS2 538–558 fragment points towards the γ-methylene of the side chain of Gln 239FIH (Fig. 5B). Previous structures have shown that Gln 239FIH binds via hydrogen-bonding interactions to the side chain of Asn residues that undergo hydroxylation, for example, Asn 803 of HIF-1α (Fig. 5C) [6,19]. However, in the TNKS2 538–558 structure, the side-chain amide of Gln 239FIH is moved away from the side chain of the hydroxylated residue (i.e. with His 553 compared with a hydroxylated Asn residue) such that it is positioned to make a hydrogen bond with the backbone amide of Tyr 102FIH. Apparently concomitant with this change, the side chain of Tyr 103FIH also moves (Fig. 5B).

Bottom Line: However, there are few reports on the selectivity of FIH for the hydroxylation of specific residues.NMR and crystallographic analyses show that the histidinyl hydroxylation occurs at the β-position.The results further expand the scope of FIH-catalysed hydroxylations.

View Article: PubMed Central - PubMed

Affiliation: Oxford Centre for Integrative Systems Biology, University of Oxford, Oxford, UK.

Show MeSH
Related in: MedlinePlus