CDNA CLONING AND OVEREXPRESSION OF 3-OXOACYL-ACP REDUCTASE FROM BRASSICA-NAPUS SEED

cDNA clones encoding NADPH-linked 3 oxoacyl-ACP reductase (EC 1.1.1.10 0) Were isolated from a Brassica napus (rape) developing seed library using a probe derived from a full length cDNA clone from Arabidopsis t haliana leaf. There is Strong sequence homology, both at the nucleotid e and amino acid level, for the cDNAs coding for this enzyme from Bras sica napus seed and Arabidopsis thaliana leaf. Northern blots analysis was performed using mRNA isolated from rape leaves and seeds, and dem onstrated that the expression of this gene in seeds is aproximately 20 times higher than in leaves. Expression of this gene in an appropriat e E coli expression vector resulted in the accumulation of the 3-oxoac yl-ACP reductase protein, which was accompanied by an increase in the enzymatic activity. The overexpressed protein was thus soluble and bio logically active. Antibodies against avocado beta-keto reductase enzym e have been previously reported to be non-cross reactive with the rape protein. On western blot of crude rape seed, no positive signal is ob served using the rabbit anti avocado antibody; however the same antibo dy strongly recognized the overproduced rape beta-keto reductase in E. coli. probably as a result of a higher concentration of the rape prot ein.