Immobilization of Candida antarctica lipase B on ILs modified CNTs with different chain lengths: Regulation of substrate tunnel “Leucine gating”

Ionic liquids (ILs) have been widely used as chemical modifiers to modify the carriers and thus improve the efficiency, activity and stability of the enzymes. However, as thousands of ILs have been found up to date, it's a huge work for screening and designing suitable ILs for immobilization of enzymes. Moreover, the mechanism of improving enzymes catalytic performance is still remain ambiguous. Thus, this study investigated the impact of ILs with different chain lengths on the enzymatic properties of Candida antarctica lipase B (CALB). Molecular dynamics simulations were employed to examine the interaction between ILs modified CNTs and CALB, as well as their effects on CALB's structure. The results revealed that ILs with different chain lengths significantly influenced the absorption orientation of CALB. Tunnel analysis identified a key role for Leu278 in regulating the open or closed state of Tunnel 2 during CALB's catalytic cycle. The weak interaction analysis demonstrated that ILs with suitable chain lengths provided spatial freedom and formed strong interactions with CNTs and ILs (vdW and hbond). This led to a conformational flip of Leu278, stabilizing the open state of Tunnel 2 and improving the activity and stability of immobilized CALB. This study provides novel insights into the design of new green modifiers to modulate carrier performance and obtain immobilized enzymes with better performance, and establishes a theoretical basis for the design and selection of modifiers for ILs in future work.