title: Enhanced thermal stability of Pseudomonas aeruginosa lipoxygenase through modification of two highly flexible regions.
abstract: Lipoxygenase (LOX; EC 1.13.11.12) is an enzyme which is widely used in food industry to improve aroma and rheological or baking properties of foods. A series of studies have proven that the flexible regions negatively relates to the thermal stability of enzymes. In this study, two highly flexible regions, residues(20-49) and residues(201-206), were modified to improve the thermal stability of LOX from Pseudomonas aeruginosa. Deletion of the first 20 and 30 residues of the former region increased the thermal stability of the LOX by 1.3- and 2.1-fold, respectively. Although deletion of the residues(201-206) led to a sharp reduction of both thermal stability and catalytic activity of the enzyme, the residue substitutions with the glycines (G204P, G206P, and G204P/G206P) or even glycine-rich linker (L6/PT) within this region increased the thermal stability of LOX by values ranging from 0.46- to 3.45-fold. To be noted, over 85% of the specific activity was maintained in all thermally stabilized LOX mutants. Circular dichroism and fluorescence analysis showed that the overall secondary and tertiary structures were not significantly changed by these modifications. To the best of our knowledge, this is the first report on increasing the thermal stability of LOX by protein engineering without remarkably affecting the catalytic rate.
authors.affiliation: Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi, 214122, China.
authors.forename: Xinyao
authors.initials: X
authors.lastname: Lu
authors.forename: Song
authors.initials: S
authors.lastname: Liu
authors.forename: Yue
authors.initials: Y
authors.lastname: Feng
authors.forename: Shengqi
authors.initials: S
authors.lastname: Rao
authors.forename: Xiaoman
authors.initials: X
authors.lastname: Zhou
authors.forename: Miao
authors.initials: M
authors.lastname: Wang
authors.forename: Guocheng
authors.initials: G
authors.lastname: Du
authors.forename: Jian
authors.initials: J
authors.lastname: Chen
mesh_terms: D002942:Circular Dichroism; D004795:Enzyme Stability; D006358:Hot Temperature; D008084:Lipoxygenase / Q000737:chemistry / Q000378:metabolism*; D011550:Pseudomonas aeruginosa / Q000201:enzymology*
chemical_list: D008084:Lipoxygenase