Identification and characterization of an arachidonate 11R-lipoxygenase

11R-Lipoxygenase (11R-LOX) activity has been detected in several marine invertebrates, and here we report the first cloning and expression of the enzyme. The cDNA encoding a protein of 77 kDa was isolated by RT-PCR from the soft coral Gersemia fruticosa and expressed in Escherichia coli. Incubations of recombinant enzyme with arachidonic acid yielded a single product, identified by RP-HPLC, GC–MS, and chiral phase-HPLC as 11R-hydroperoxyeicosatetraenoic acid. Other C18, C20, and C22 substrates are also oxygenated, preferentially at the ω10 position. Significantly, both Ca2+-ions and a membrane fraction are required for catalytic activity. Calcium effects translocation of the soluble 11R-LOX to the membrane and this association is reversible by Ca2+ chelation. The enzyme sequence contains some conserved amino acids implicated in calcium activation of mammalian 5-LOX, and with its obligate requirement for membrane interaction the 11R-LOX may thus provide a new model for further analysis of this aspect of lipoxygenase activation.