AN OLEATE 12-HYDROXYLASE FROM RICINUS-COMMUNIS L IS A FATTY ACYL DESATURASE HOMOLOG

Recent spectroscopic evidence implicating a binuclear iron site at the reaction center of fatty acyl desaturases suggested to us that certai n fatty acyl hydroxylases may share significant amino acid sequence si milarity with desaturases. To test this theory, we prepared a cDNA lib rary from developing endosperm of the castor-oil plant (Ricinus commun is L.) and obtained partial nucleotide sequences for 468 anonymous clo nes that were not expressed at high levels in leaves, a tissue deficie nt in 12-hydroxyoleic acid. This resulted in the identification of sev eral cDNA clones encoding a polypeptide of 387 amino acids with a pred icted molecular weight of 44,407 and with approximate to 67% sequence homology to microsomal oleate desaturase from Arabidopsis. Expression of a full-length clone under control of the cauliflower mosaic virus 3 5S promoter in transgenic tobacco resulted in the accumulation of low levels of 12-hydroxyoleic acid in seeds, indicating that the clone enc odes the castor oleate hydroxylase. These results suggest that fatty a cyl desaturases and hydroxylases share similar reaction mechanisms and provide an example of enzyme evolution.