Biogenesis of volatile aldehydes from fatty acid hydroperoxides: molecular cloning of a hydroperoxide lyase (CYP74C) with specificity for both the 9- and 13-hydroperoxides of linoleic and linolenic acids.

Tijet N, Schneider C, Muller BL, Brash AR
Arch Biochem Biophys. 2001 386 (2): 281-9

PMID: 11368353 · DOI:10.1006/abbi.2000.2218

A novel member of the plant cytochrome P450 CYP74 family of fatty acid hydroperoxide metabolizing enzymes has been cloned from melon fruit (Cucumis melo). The cDNA is comprised of 1,446 nucleotides encoding a protein of 481 amino acids. The homology at the amino acid level to other members of the CYP74 family is 35-50%, the closest relatives being allene oxide synthases. The cDNA was expressed in Escherichia coli, and the corresponding protein was purified by affinity column chromatography. The native enzyme showed a main Soret band at 418 nm, indicative of a low spin ferric cytochrome P450, and a 447-nm peak appeared in the CO-difference spectrum. Using [U-14C]radiolabeled substrate, HPLC, UV, and GC-MS, the products of conversion of 9S-hydroperoxy-linoleic acid were identified as 9-oxo-nonanic acid and 3Z-nonenal. Kinetic analysis of this hydroperoxide lyase showed the highest rate of reaction with 9-hydroperoxy-linolenic acid followed by 9-hydroperoxy-linoleic acid and then the corresponding 13-hydroperoxides. Overall, the newly characterized cytochrome P450 enzyme is a fatty acid hydroperoxide lyase with a preference, but not absolute specificity for the 9-positional hydroperoxides of linoleic and linolenic acids.

MeSH Terms (21)

Aldehyde-Lyases Aldehydes alpha-Linolenic Acid Amino Acid Sequence Base Sequence Catalysis Chromatography, High Pressure Liquid Cloning, Molecular Cucurbitaceae Cytochrome P-450 Enzyme System Escherichia coli Gas Chromatography-Mass Spectrometry Hydrogen-Ion Concentration Kinetics Linoleic Acid Lipid Peroxides Molecular Sequence Data Phylogeny Recombinant Fusion Proteins Spectrum Analysis Substrate Specificity

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