Creating lipoxygenases with new positional specificities by site-directed mutagenesis. (December 2000)

• Record Type: Journal Article
• Authors: Hornung, E.; Rosahl, S.; Kühn, H.; Feussner, I.
• Is Part Of: Biochemical Society transactions. Volume 28:Number 6(2000)
• Page Start: 825
• Page End: 826
• Publication Date: 2000-12
• Subjects: active site -- enzyme engineering -- hydroperoxy polyenoic fatty acid; LOX, lipoxygenase; Biochemistry -- Congresses; 572
• DOI: 10.1042/bst0280825
• Languages: English
• ISSNs: 0300-5127

Abstract

Abstract : In order to analyse the amino acid determinants which alter the positional specificity of plant lipoxygenases (LOXs), multiple LOX sequence alignments and structural modelling of the enzyme-substrate interactions were carried out. These alignments suggested three amino acid residues as the primary determinants of positional specificity. Here we show the generation of two plant LOXs with new positional specificities, a Δ-linoleneate 6-LOX and an arachidonate 11-LOX, by altering only one of these determinants within the active site of two plant LOXs. In the past, site-directed-mutagenesis studies have mainly been carried out with mammalian lipoxygenases (LOXs) [1]. In these experiments two regions have been identified in the primary structure containing sequence determinants for positional specificity. Amino acids aligning with the Sloane determinants [2] are highly conserved among plant LOXs. In contrast, there is amino acid hetero-geneity among plant LOXs at the position that aligns with P353 of the rabbit reticulocyte 15-LOX (Borngräber determinants) [3].