Dataset Information

Biochemical and Structural Insights into FIH-Catalysed Hydroxylation of Transient Receptor Potential Ankyrin Repeat Domains.

ABSTRACT: Transient receptor potential (TRP) channels have important roles in environmental sensing in animals. Human TRP subfamily A member 1 (TRPA1) is responsible for sensing allyl isothiocyanate (AITC) and other electrophilic sensory irritants. TRP subfamily vanilloid member 3 (TRPV3) is involved in skin maintenance. TRPV3 is a reported substrate of the 2-oxoglutarate oxygenase factor inhibiting hypoxia-inducible factor (FIH). We report biochemical and structural studies concerning asparaginyl hydroxylation of the ankyrin repeat domains (ARDs) of TRPA1 and TRPV3 catalysed by FIH. The results with ARD peptides support a previous report on FIH-catalysed TRPV3 hydroxylation and show that, of the 12 potential TRPA1 sequences investigated, one sequence (TRPA1 residues 322-348) undergoes hydroxylation at Asn336. Structural studies reveal that the TRPA1 and TRPV3 ARDs bind to FIH with a similar overall geometry to most other reported FIH substrates. However, the binding mode of TRPV3 to FIH is distinct from that of other substrates.

SUBMITTER: Saward BG

PROVIDER: S-EPMC10946520 | biostudies-literature | 2023 Feb

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Biochemical and Structural Insights into FIH-Catalysed Hydroxylation of Transient Receptor Potential Ankyrin Repeat Domains.

Saward Benjamin G BG Leissing Thomas M TM Clifton Ian J IJ Tumber Anthony A Timperley Christopher M CM Hopkinson Richard J RJ Schofield Christopher J CJ

Chembiochem : a European journal of chemical biology 20230109 4

Transient receptor potential (TRP) channels have important roles in environmental sensing in animals. Human TRP subfamily A member 1 (TRPA1) is responsible for sensing allyl isothiocyanate (AITC) and other electrophilic sensory irritants. TRP subfamily vanilloid member 3 (TRPV3) is involved in skin maintenance. TRPV3 is a reported substrate of the 2-oxoglutarate oxygenase factor inhibiting hypoxia-inducible factor (FIH). We report biochemical and structural studies concerning asparaginyl hydroxy ...[more]

PMID: 36448355

Similar Datasets

Project description:The factor inhibiting HIF (FIH) is a proximate oxygen sensor for human cells, hydroxylating Asn(803) within the ?-subunit of the hypoxia inducible factor (HIF). FIH is an ?-ketoglutatrate (?KG)-dependent, non-heme Fe(II) dioxygenase, in which Fe(II) is coordinated by a (His(2)Asp) facial triad, ?KG, and H(2)O. Hydrogen bonding among the facial triad, the HIF-Asn(803) side chain, and various second-sphere residues suggests a functional role for the second coordination sphere in tuning the chemistry of the Fe(II) center. Point mutants of FIH were prepared to test the functional role of the ?KG-centered (Asn(205) and Asn(294)) or HIF-Asn(803)-centered (Arg(238) and Gln(239)) second-sphere residues. The second sphere was tested for local effects on priming Fe(II) to react with O(2), oxidative decarboxylation, and substrate positioning. Steady-sate kinetics were used to test for overall catalytic effects; autohydroxylation rates were used to test for priming and positioning, and electronic spectroscopy was used to assess the primary coordination sphere and the electrophilicity of ?KG. Asn(205) ? Ala and Asn(294) ? Ala mutants exhibited diminished rates of steady-state turnover, while minimally affecting autohydroxylation, consistent with impaired oxidative decarboxylation. Blue-shifted metal to ligand charge transfer transitions for (Fe+?KG)FIH indicated that these point mutations destabilized the ?* orbitals of ?KG, further supporting a slowed rate of oxidative decarboxylation. The Arg(238) ? Met mutant exhibited steady-state rates too low to measure and diminished product yields, suggesting impaired substrate positioning or priming; the Arg(238) ? Met mutant was capable of O(2) activation for the autohydroxylation reaction. The Gln(239) ? Asn mutant exhibited significantly slowed steady-state kinetics and diminished product yields, suggesting impaired substrate positioning or priming. As HIF binding to the Gln(239) ? Asn mutant stimulated autohydroxylation, it is more likely that this point mutant simply mispositions the HIF-Asn(803) side chain. This work combines kinetics and spectroscopy to show that these second-sphere hydrogen bonds play roles in promoting oxidative decarboxylation, priming Fe(II) to bind O(2), and positioning HIF-Asn(803).

Dataset Information

Biochemical and Structural Insights into FIH-Catalysed Hydroxylation of Transient Receptor Potential Ankyrin Repeat Domains.

Publications

Biochemical and Structural Insights into FIH-Catalysed Hydroxylation of Transient Receptor Potential Ankyrin Repeat Domains.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets