Studies on the specificity of unprocessed and mature forms of phytanoyl-CoA 2-hydroxylase and mutation of the iron binding ligands.
The mature form of phytanoyl-coenzyme A 2-hydroxylase (PAHX), a nonheme Fe(II)- and 2-oxoglutarate-dependent oxygenase, catalyzes the alpha-hydroxylation of phytanoyl-CoA within peroxisomes. Mutations in PAHX result in some forms of adult Refsum's disease. Unprocessed PAHX (pro-PAHX) contains a...
| Main Authors: | , , , , , |
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| Format: | Journal article |
| Language: | English |
| Published: |
2005
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| _version_ | 1797105970653954048 |
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| author | Searls, T Butler, D Chien, W Mukherji, M Lloyd, MD Schofield, C |
| author_facet | Searls, T Butler, D Chien, W Mukherji, M Lloyd, MD Schofield, C |
| author_sort | Searls, T |
| collection | OXFORD |
| description | The mature form of phytanoyl-coenzyme A 2-hydroxylase (PAHX), a nonheme Fe(II)- and 2-oxoglutarate-dependent oxygenase, catalyzes the alpha-hydroxylation of phytanoyl-CoA within peroxisomes. Mutations in PAHX result in some forms of adult Refsum's disease. Unprocessed PAHX (pro-PAHX) contains an N-terminal peroxisomal targeting sequence that is cleaved to give mature PAHX (mat-PAHX). Previous studies have implied a difference in the substrate specificity of the unprocessed and mature forms of PAHX. We demonstrate that both forms are able to hydroxylate a range of CoA derivatives, but under the same assay conditions, the N-terminal hexa-His-tagged unprocessed form is less active than the nontagged mature form. Analyses of the assay conditions suggest a rationale for the lack of activity previously reported for some substrates (e.g. isovaleryl-CoA) for the (His)6pro-PAHX. Site-directed mutagenesis was used to support proposals for the identity of the iron binding ligands (His-175, Asp-177, His-264) of the 2-His-1-carboxylate motif of PAHX. Mutation of other histidine residues (His-213, His-220, His-259) suggested that these residues were not involved in Fe(II) binding. |
| first_indexed | 2024-03-07T06:55:00Z |
| format | Journal article |
| id | oxford-uuid:fdd60a79-5773-475f-8efb-d84304a0ec37 |
| institution | University of Oxford |
| language | English |
| last_indexed | 2024-03-07T06:55:00Z |
| publishDate | 2005 |
| record_format | dspace |
| spelling | oxford-uuid:fdd60a79-5773-475f-8efb-d84304a0ec372022-03-27T13:31:45ZStudies on the specificity of unprocessed and mature forms of phytanoyl-CoA 2-hydroxylase and mutation of the iron binding ligands.Journal articlehttp://purl.org/coar/resource_type/c_dcae04bcuuid:fdd60a79-5773-475f-8efb-d84304a0ec37EnglishSymplectic Elements at Oxford2005Searls, TButler, DChien, WMukherji, MLloyd, MDSchofield, CThe mature form of phytanoyl-coenzyme A 2-hydroxylase (PAHX), a nonheme Fe(II)- and 2-oxoglutarate-dependent oxygenase, catalyzes the alpha-hydroxylation of phytanoyl-CoA within peroxisomes. Mutations in PAHX result in some forms of adult Refsum's disease. Unprocessed PAHX (pro-PAHX) contains an N-terminal peroxisomal targeting sequence that is cleaved to give mature PAHX (mat-PAHX). Previous studies have implied a difference in the substrate specificity of the unprocessed and mature forms of PAHX. We demonstrate that both forms are able to hydroxylate a range of CoA derivatives, but under the same assay conditions, the N-terminal hexa-His-tagged unprocessed form is less active than the nontagged mature form. Analyses of the assay conditions suggest a rationale for the lack of activity previously reported for some substrates (e.g. isovaleryl-CoA) for the (His)6pro-PAHX. Site-directed mutagenesis was used to support proposals for the identity of the iron binding ligands (His-175, Asp-177, His-264) of the 2-His-1-carboxylate motif of PAHX. Mutation of other histidine residues (His-213, His-220, His-259) suggested that these residues were not involved in Fe(II) binding. |
| spellingShingle | Searls, T Butler, D Chien, W Mukherji, M Lloyd, MD Schofield, C Studies on the specificity of unprocessed and mature forms of phytanoyl-CoA 2-hydroxylase and mutation of the iron binding ligands. |
| title | Studies on the specificity of unprocessed and mature forms of phytanoyl-CoA 2-hydroxylase and mutation of the iron binding ligands. |
| title_full | Studies on the specificity of unprocessed and mature forms of phytanoyl-CoA 2-hydroxylase and mutation of the iron binding ligands. |
| title_fullStr | Studies on the specificity of unprocessed and mature forms of phytanoyl-CoA 2-hydroxylase and mutation of the iron binding ligands. |
| title_full_unstemmed | Studies on the specificity of unprocessed and mature forms of phytanoyl-CoA 2-hydroxylase and mutation of the iron binding ligands. |
| title_short | Studies on the specificity of unprocessed and mature forms of phytanoyl-CoA 2-hydroxylase and mutation of the iron binding ligands. |
| title_sort | studies on the specificity of unprocessed and mature forms of phytanoyl coa 2 hydroxylase and mutation of the iron binding ligands |
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