共 25 条
Structural basis for pH-dependent alterations of reaction specificity of vertebrate lipoxygenase isoforms
被引:18
作者:
Walther, M.
[1
]
Roffeis, J.
[1
]
Jansen, C.
[1
]
Anton, M.
[1
]
Ivanov, I.
[1
]
Kuhn, H.
[1
]
机构:
[1] Univ Med Berlin, Charite, Inst Biochem, D-10117 Berlin, Germany
来源:
BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR AND CELL BIOLOGY OF LIPIDS
|
2009年
/
1791卷
/
08期
关键词:
Eicosanoids;
Lipoxygenase;
Fatty acid metabolism;
Molecular enzymology;
Catalytic mechanism;
POSITIONAL SPECIFICITY;
ACTIVE-SITE;
ACID;
5-LIPOXYGENASE;
DETERMINANTS;
CONVERSION;
IRON;
8S-LIPOXYGENASE;
IDENTIFICATION;
OXYGENATION;
D O I:
10.1016/j.bbalip.2009.05.007
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:
Lipoxygenases have been classified according to their specificity of fatty acid oxygenation and for several plant enzymes pH-dependent alterations in the product patterns have been reported. Assuming that the biological role of mammalian lipoxygenases is based on the formation of specific reaction products, pH-dependent alterations would impact enzymes' functionality. In this study we systematically investigated the pH-dependence of vertebrate lipoxygenases and observed a remarkable stability of the product pattern in the near physiological range for the wild-rype enzyme species. Site-directed mutagenesis of selected amino acids and alterations in the substrate concentrations induced a more pronounced pH-dependence of the reaction specificity. For instance, for the V603H mutant of the human 15-lipoxygenase-2 8-lipoxygenation was dominant at acidic pH (65%) whereas 15-H(p)ETE was the major oxygenation product at pH 8. Similarly, the product pattern of the wild-type mouse 8-lipoxygenase was hardly altered in the near physiological pH range but H604F exchange induced strong pH-dependent alterations in the positional specificity. Taken together, our data suggest that the reaction specificities of wild-type vertebrate lipoxygenase isoforms are largely resistant towards pH alterations. However, we found that changes in the assay conditions (low substrate concentration) and introduction/removal of a critical histidine at the active site impact the pH-dependence of reaction specificity for some lipoxygenase isoforms. (C) 2009 Elsevier B.V. All rights reserved.
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页码:827 / 835
页数:9
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