Enhanced Thermostability of Lipoxygenase from Anabaena sp. PCC 7120 by Site-Directed Mutagenesis Based on Computer-Aided Rational Design.

Diao, Hanwen; Zhang, Chong; Wang, Shuicheng; Lu, Fengxia; Lu, Zhaoxin

Lipoxygenase from Anabaena sp. PCC 7120 (Ana-LOX) was thermally unstable. So, improving the thermostability of the enzyme was quite essential. The target site of Ana-LOX selected for site-directed mutagenesis was based on computer-aided rational design. The thermostability and specific activity of Ana-LOX were improved with replacing valine with alanine at the target site 421 and the site 40. Compared to the wild-type enzyme which has a half-life ( T) of inactivation of 3.8 min at 50 °C, the T of mutant enzymes with V421A and V40A substitution increased to 4.4 and 7.0 min, respectively. The double mutant V421A/V40A showed a synergistic effect with a T value of 8.3 min, resulting in a 1.18-fold improvement compared to the original Ana-LOX. V421A, V40A, and V421A/V40A also obtained 4.83, 41.58, and 80.07 % increase in specific activity, respectively. This study provides useful theoretical reference for enzyme molecular modification and computer-aided rational design.

LIPOXYGENASES; OXYGENASES; ARACHIDONATE 12-lipoxygenase; ANABAENA; NOSTOCACEAE

Applied Biochemistry & Biotechnology, 2016, Vol 178, Issue 7, p1339

0273-2289

Academic Journal

10.1007/s12010-015-1950-2