Site‐directed mutagenesis studies of hydrophobic residues in the lid region of T1 lipase


T1 lipase is a potential biocatalyst for industrial application since it is highly thermostable and displays optimal activity at 60–75°C. Structural analysis of T1 lipase shows that its lid region undergoes a spatial displacement along with a distinct secondary structure reorganization upon activation. To study structure/function of this atypical lid, we performed site‐directed mutagenesis on the hydrophobic residues in the lid region. These residues were mutated to hydrophilic ones and biochemical properties of mutants were investigated. Results showed that F181 might be an important residue for enzyme‐substrate binding. Mutants A186S and A190S had 35–50% increase in catalytic efficiencies compared to wild‐type T1, without compromising their functions at high temperatures. In general, mutagenesis did not cause large changes to chain‐length preference in T1 lipase. Mutants A186S and V187N were inactive towards long‐chain pNP esters (p‐Nitrophenyl stearate) and V187N showed lower activity towards long‐chain triacylglycerols than wt T1, which makes it a potential catalyst in dairy industry. Thermostability of mutants were affected at different extent due to the influence of hydrophobic contact between the lid and the protein core. These findings not only shed light on the lipase structure/function relationship but also lay the framework for further engineering to gain more potent, stable, and selective lipases.

Submission Details

ID: xL7zWse23

Submitter: Shu-Ching Ou

Submission Date: Oct. 11, 2018, 2:13 p.m.

Version: 1

Publication Details
Tang Q;Lan D;Yang B;Khan FI;Wang Y,Eur. J. Lipid Sci. Technol. (2016) Site‐directed mutagenesis studies of hydrophobic residues in the lid region of T1 lipase
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