Lipases are important industrial enzymes. Most of the lipases operate at lipid-water interfaces enabled by a mobile lid domain located over the active site. Lid protects the active site and hence responsible for catalytic activity. In pure aqueous media the lid is predominantly closed, whereas in the presence of a hydrophobic layer it is partially opened. Hence, the lid controls the enzyme activity. In the present review, we have classified lipases into different groups based on the structure of lid domains. It has been observed that thermostable lipases contain larger lid domains with two or more helices, whereas mesophilic lipases tend to have smaller lids in the form of a loop or a helix. Recent developments in lipase engineering addressing the lid regions are critically reviewed here. After on the dramatic changes in substrate selectivity, activity and thermostability have been reported. Furthermore, improved computational models now can rationalize these observations by relating it to the mobility of the lid domain. In this contribution, we summarized and critically evaluated the most recent developments in experimental and computational research on lipase lids.

Lipases with unique substrate specificity are highly desired in biotechnological applications. In this study, a putative marine Geobacillus sp. monoacylglycerol lipase (GMGL) encoded gene was identified by a genomic mining strategy. The gene was expressed in Escherichia coli as a His-tag fusion protein and purified by affinity chromatography with a yield of 264 mg per liter fermentation broth. The recombinant GMGL shows the highest hydrolysis activity at 60 °C and pH 8.0, and the half-life was 60 min at 70 °C. The GMGL is active on monoacylglycerol (MAG) substrate but not diacylglycerol (DAG) or triacylglycerol (TAG), and produces MAG as the single product in the esterification reaction. Modeling structure analysis showed that the catalytic triad is formed by Ser97, Asp196 and His226, and the flexible cap region is constituted by residues from Ala120 to Thr160. A mutagenesis study on Leu142, Ile145 and Ile170 located in the substrate binding tunnel revealed that these residues were related with its substrate specificity. The k_cat/K_m value toward the pNP-C6 substrate in mutants Leu142Ala, Ile145Ala and Ile170Phe increased to 2.3-, 1.4- and 2.2-fold as compared to that of the wild type, respectively.

Two effective molecularly imprinted polymers for the adsorption of alpha-lipoic acid (ALA) were synthesized by the cross-linking of chitosan with epichlorohydrin (ECH) and glutaraldehyde (GLU), respectively, in the presence of ALA as template molecules. Investigations on the molar ratios of ALA and chitosan (–NH₂) in the preparation of chitosan molecularly imprinted polymers (MIPs) were carried out with a factor of ALA rebinding capabilities. The surface morphology and chemical properties of the polymers were characterized. The optimized MIPs crosslinked by ECH (MIPs–ECH) and MIPs crosslinked by GLU (MIPs–GLU) had adsorption capabilities of 12.09 mg/g and 19.72 mg/g for ALA, respectively. The adsorption behaviors of two kinds of chitosan MIPs including adsorption kinetics and isotherms were investigated in detail. Adsorption and kinetic binding experiments showed that the prepared MIPs–ECH and MIPs–GLU had selective adsorption and excellent affinity for ALA. In addition, the possible binding models between ALA and chitosan oligosaccharide were predicted by molecular dynamics simulation.