Structure-function analysis of phytanoyl-CoA 2-hydroxylase mutations causing Refsum's disease

被引:53
作者
Mukherji, M
Chien, W
Kershaw, NJ
Clifton, IJ
Schofield, CJ
Wierzbicki, AS
Lloyd, MD
机构
[1] Oxford Ctr Mol Sci, Oxford OX1 3QY, England
[2] Dyson Perrins Lab, Oxford OX1 3QY, England
[3] St Thomas Hosp, Dept Chem Pathol, Kings Guys & St Thomas Med Sch, London SE1 7EH, England
关键词
D O I
10.1093/hmg/10.18.1971
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Refsum's disease is a neurological syndrome characterized by adult-onset retinitis pigmentosa, anosmia, sensory neuropathy and phytanic acidaemia. Many cases are caused by mutations in peroxisomal oxygenase phytanoyl-CoA 2-hydroxylase (PAHX) which catalyses the initial alpha-oxidation step in the degradation of phytanic acid. Both pro and mature forms of recombinant PAHX were produced in Escherichia coli, highly purified, and shown to have a requirement for iron(II) as a co-factor and 2-oxoglutarate as a co-substrate. Sequence analysis in the light of crystallographic data for other members of the 2-oxoglutarate-dependent oxygenase super-family led to secondary structural predictions for PAHX, which were tested by site-directed mutagenesis. The H175A and D177A mutants did not catalyse hydroxylation of phytanoyl-CoA, consistent with their assigned role as iron(II) binding ligands. The clinically observed P29S, Q176K, G204S, N269H, R275Q and R275W mutants were assayed for both 2-oxoglutarate and phytanoyl-CoA oxidation. The P29S mutant was fully active, implying that the mutation resulted in defective targeting of the protein to peroxisomes. Mutation of Arg-275 resulted in impaired 2-oxoglutarate binding. The Q176K, G204S and N269H mutations caused partial uncoupling of 2-oxoglutarate conversion from phytanoyl-CoA oxidation. The results demonstrate that the diagnosis of Refsum's disease should not solely rely upon PAHX assays for 2-oxoglutarate or phytanoyl-CoA oxidation.
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页码:1971 / 1982
页数:12
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共 56 条
  • [1] Aravind L, 2001, GENOME BIOL, V2
  • [2] ALSCRIPT - A TOOL TO FORMAT MULTIPLE SEQUENCE ALIGNMENTS
    BARTON, GJ
    [J]. PROTEIN ENGINEERING, 1993, 6 (01): : 37 - 40
  • [3] BRADFORD MM, 1976, ANAL BIOCHEM, V72, P248, DOI 10.1016/0003-2697(76)90527-3
  • [4] Croes K, 2000, J LIPID RES, V41, P629
  • [5] Croes K, 1999, J LIPID RES, V40, P601
  • [6] Production of formyl-CoA during peroxisomal alpha-oxidation of 3-methyl-branched fatty acids
    Croes, K
    VanVeldhoven, PP
    Mannaerts, GP
    Casteels, M
    [J]. FEBS LETTERS, 1997, 407 (02) : 197 - 200
  • [7] NOVEL INHIBITORS OF PROLYL 4-HYDROXYLASE .3. INHIBITION BY THE SUBSTRATE-ANALOG N-OXALOGLYCINE AND ITS DERIVATIVES
    CUNLIFFE, CJ
    FRANKLIN, TJ
    HALES, NJ
    HILL, GB
    [J]. JOURNAL OF MEDICINAL CHEMISTRY, 1992, 35 (14) : 2652 - 2658
  • [8] Alkyl-dihydroxyacetonephosphate synthase - Fate in peroxisome biogenesis disorders and identification of the point mutation underlying a single enzyme deficiency
    de Vet, ECJM
    Ijlst, L
    Oostheim, W
    Wanders, RJA
    van den Bosch, H
    [J]. JOURNAL OF BIOLOGICAL CHEMISTRY, 1998, 273 (17) : 10296 - 10301
  • [9] Mutagenesis studies on the iron binding ligands of clavaminic acid synthase
    Doan, LX
    Hassan, A
    Lipscomb, SJ
    Dhanda, A
    Zhang, ZH
    Schofield, CJ
    [J]. BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 2000, 279 (01) : 240 - 244
  • [10] Probing the penicillin sidechain selectivity of recombinant deacetoxycephalosporin C synthase
    Dubus, A
    Lloyd, MD
    Lee, HJ
    Schofield, CJ
    Baldwin, JE
    Frère, JM
    [J]. CELLULAR AND MOLECULAR LIFE SCIENCES, 2001, 58 (5-6) : 835 - 843