Properties and structural requirements for substrate specificity of cytochrome P-450-dependent obtusifoliol 14 α-demethylase from maize (Zea mays) seedlings
| Autor: | M Taton, A Rahier |
|---|---|
| Rok vydání: | 1991 |
| Předmět: |
Cytochrome
Stereochemistry Molecular Conformation Zea mays Biochemistry Substrate Specificity Sterol 14-Demethylase chemistry.chemical_compound Cytochrome P-450 Enzyme System Microsomes Binding site Molecular Biology Demethylation chemistry.chemical_classification Molecular Structure biology Lanosterol Cholestadienols Phytosterols Cytochrome P450 Substrate (chemistry) Cell Biology Kinetics Enzyme chemistry biology.protein Oxidoreductases Research Article |
| Zdroj: | Biochemical Journal. 277:483-492 |
| ISSN: | 1470-8728 0264-6021 |
| DOI: | 10.1042/bj2770483 |
| Popis: | The biochemical properties of cytochrome P-450-dependent obtusifoliol 14 alpha-demthylase (P-450OBT.14DM) from maize (Zea mays) seedlings were defined. In particular, the enzyme was shown by differential centrifugation to be localized in the endoplasmic reticulum. P-450OBT.14DM had an apparent Km of 160 +/- 5 microM and an apparent Vmax of 65 +/- 5 pmol/min per mg of protein for its best substrate, obtusifoliol. The substrate specificity of P-450OBT.14DM was thoroughly investigated by comparing the demethylation of obtusifoliol with that of a series of 15 natural or novel synthetic analogues of obtusifoliol. The results obtained clearly indicate that three distinct domains of the sterol substrate are governing obtusifoliol demethylation by P-450OBT.14DM. They revealed that (i) P-450OBT.14DM has probably a specific apolar binding site for the side chain, (ii) the delta 8-double bond is an absolute requirement for substrate demethylation and (iii) the 3-hydroxy group plays a critical role in the enzyme-substrate interaction. Interestingly the binding site, beyond the C-3 position, contains a cleft which cannot accommodate a 4 beta-methyl substituent present in lanosterol or eburicol, the precursors of 14-desmethylsterols respectively in mammals and yeast. This result indicates that P-450OBT.14DM is a novel constitutive cytochrome P-450 with a high degree of substrate and product specificity. |
| Databáze: | OpenAIRE |
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