Four major enzymes commonly used in the market are lipases, proteases, amylases, and cellulases. For instance, in both academic and industrial levels, microbial lipases have been well studied for industrial and biotechnological applications compared to others. Immobilization is done to minimize the cost. The improvement of enzyme properties enables the reusability of enzymes and facilitates enzymes used in a continuous process. Immobilized enzymes are enzymes physically confined in a particularly defined region with retention to their catalytic activities. Immobilized enzymes can be used repeatedly compared to free enzymes, which are unable to catalyze reactions continuously in the system. Immobilization also provides a higher pH value and thermal stability for enzymes toward synthesis. The main parameter influencing the immobilization is the support used to immobilize the enzyme. The support should have a large surface area, high rigidity, suitable shape and particle size, reusability, and resistance to microbial attachment, which will enhance the stability of the enzyme. The diffusion of the substrate in the carrier is more favorable on hydrophobic supports instead of hydrophilic supports. The methods used for enzyme immobilization also play a crucial role in immobilization performance. The combination of immobilization methods will increase the binding force between enzymes and the support, thus reducing the leakage of the enzymes from the support. The adsorption of lipase on a hydrophobic support causes the interfacial activation of lipase during immobilization. The adsorption method also causes less or no change in enzyme conformation, especially on the active site of the enzyme. Thus, this method is the most used in the immobilization process for industrial applications.

Microbial lipases represent one of the most important groups of biotechnological biocatalysts. However, the high-level production of lipases requires an understanding of the molecular mechanisms of gene expression, folding, and secretion processes. Stable, selective, and productive lipase is essential for modern chemical industries, as most lipases cannot work in different process conditions. However, the screening and isolation of a new lipase with desired and specific properties would be time consuming, and costly, so researchers typically modify an available lipase with a certain potential for minimizing cost. Improving enzyme properties is associated with altering the enzymatic structure by changing one or several amino acids in the protein sequence. This review detailed the main sources, classification, structural properties, and mutagenic approaches, such as rational design (site direct mutagenesis, iterative saturation mutagenesis) and direct evolution (error prone PCR, DNA shuffling), for achieving modification goals. Here, both techniques were reviewed, with different results for lipase engineering, with a particular focus on improving or changing lipase specificity. Changing the amino acid sequences of the binding pocket or lid region of the lipase led to remarkable enzyme substrate specificity and enantioselectivity improvement. Site-directed mutagenesis is one of the appropriate methods to alter the enzyme sequence, as compared to random mutagenesis, such as error-prone PCR. This contribution has summarized and evaluated several experimental studies on modifying the substrate specificity of lipases.

In recent years, studies on psychrophilic lipases have been an emerging area of research in the field of enzymology. This study focuses on bacterial strains isolated from anthropogenically-influenced soil samples collected around Signy Island Research Station (South Orkney Islands, maritime Antarctic). Limited information on lipase activities from bacteria isolated from Signy station is currently available. The presence of lipase genes was determined using real time quantification PCR (qPCR) in samples obtained from three different locations on Signy Island. Twenty strains from the location with highest lipase gene detection were screened for lipolytic activities at a temperature of 4 °C, and from this one strain was selected for further examination based on the highest enzymatic activities obtained. Analysis of 16S rRNA sequence data of this strain showed the highest level of sequence similarity (98%) to a Pseudomonas sp. strain also isolated from Antarctica. In order to increase lipase production of this psychrophilic strain, optimisation of different parameters of physical and nutritional factors were investigated. Optimal production was obtained at 10 °C and pH 7.0, at 150 rev/min shaking rate over 36 h incubation.

The use of T1 lipase in automatic dishwashing detergent (ADD) is well established, but efficiency in hard water is very low. A new enzymatic environmentally-friendly dishwashing was formulated to be efficient in both soft and hard water. Thermostable enzymes such as T1 lipase from Geobacillus strain T1, Rand protease from Bacillus subtilis strain Rand, and Maltogenic amylase from Geobacillus sp. SK70 were produced and evaluated for an automatic dishwashing detergent formulation. The components of the new ADD were optimized for compatibility with these three enzymes. In compatibility tests of the enzymes with different components, several criteria were considered. The enzymes were mostly stable in non-ionic surfactants, especially polyhydric alcohols, Glucopon UP 600, and in a mixture of sodium carbonate and glycine (30:70) buffer at a pH of 9.25. Sodium polyacrylate and sodium citrate were used in the ADD formulation as a dispersing agent and a builder, respectively. Dishwashing performance of the formulated ADDs was evaluated in terms of percent of soil removed using the Leenert‘s Improved Detergency Tester. The results showed that the combination of different hydrolysis enzymes could improve the washing efficiency of formulated ADD compared to the commercial ADD “Finish” at 40 and 50 C.