Effect of cavity-filling mutations on thermostability of lipase LipA from Burkholderia sp.

Abstract:

[Objective] To improve the thermostability of lipase LipA from Burkholderia sp., a series of cavity-filled lipase LipA mutants were constructed and evaluated. [Methods] Amino acid residuals consisted of cavities in 3D structural model of lipase LipA were predicted using Castp, Voronoia and Cave, and then computational libraries of every amino acid residuals were constructed using software packages FoldX. The following types of mutants were excluded from the computational libraries:(1) volume of the cavity was increased; (2) value of free energy, ΔΔG, was over -0.5 kcal/mol. Total 58 mutants were screened from the computational libraries, and 17 mutants were selected to verify the mutation effect. [Results] Thermostability of lipase mutants (LipA-His¹⁵Pro and LipA-Ala²¹⁰Val) were improved and T₅₀¹² value of lipase LipA-His¹⁵Pro and lipase LipA-Ala²¹⁰Val increased by 4 ℃ and 2 ℃, respectively. Thermostability of the superimposing mutant lipase LipA-His¹⁵Pro/Ala²¹⁰Val was further improved and the half-life (t_1/2) at 55 ℃ increased by 23.1 times. [Conclusion] Cavity-filling mutation was a feasible technique to improve the thermostability of lipase LipA from Burkholderia sp.