The alternative pathway C20 Δ8-desaturase from the non-photo synthetic organism Acanthamoeba castellanii is an atypical cytochrome b5-fusion desaturase

A cDNA encoding a C-20 Delta 8-desaturase was isolated from the free-living soil amoeba, A canthamoeba castellanii and functionally characterised by heterologous expression. The open reading frame of the A. castellanii CM A8-desaturase showed similarity to other microsomal front-end desaturases, but the N-terminal domain contained a variant form of the conserved heme-binding motif in which H-P-G-G is replaced by H-P-A-G. Co-expression of the A. castellani Delta 8-desaturase with the Isochrysis galbana Delta 9-elongase in transgenic Arabidopsis plants confirmed the activity observed in yeast and its role in the alternative pathway for C-20 polyunsaturated fatty acid synthesis. Acyl-CoA profiles of these transgenic plants revealed an unexpected accumulation of C-20 fatty acids in the acyl-CoA pool. This is the first report of an alternative pathway C-20 Delta 8-desaturase from a non-photosynthetic organism, and also the first report of a front-end desaturase lacking the canonical cytochrome b(5) domain. (c) 2006 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.