**1. Introduction**

The industrial enzymes market is estimated to be valued at USD 5.9 billion in 2020 and is projected to reach USD 8.7 billion by 2026, recording a Compound Annual Growth Rate (CAGR) of 6.5%, in terms of value [1]. The majority of enzymes currently used in industrial processes (more than 75%) are hydrolases [2]. Lipases represent the third most commercialized enzymes, after carbohydrases and proteases [3], and their production has constantly increased, they now account for more than one-fifth of the global enzyme market. The global Lipase Market size is anticipated to develop at a notable CAGR of about 7% over the calculated period from the current value of USD 0.6 billion in 2020. Lipases form an integral part of the industries ranging from biodiesels, food, nutraceuticals and detergents with little utilization in bioremediation, agriculture, cosmetics and leather [4].

Although lipases are produced by a huge number of organisms (bacterial, plant and animal origin), microbial lipases have attracted far more interest from researchers and industries than lipases from other sources, due to both their specific features and ease of production on large scale [5–7]. Notwithstanding current achievements, there is still a quest for lipases with improved and/or novel catalytic features like stability in harsh environments. Marine organisms can be an adequate source for such lipases

**Citation:** Ben Hlima, H.; Dammak, M.; Karray, A.; Drira, M.; Michaud, P.; Fendri, I.; Abdelkafi, S. Molecular and Structural Characterizations of Lipases from *Chlorella* by Functional Genomics. *Mar. Drugs* **2021**, *19*, 70. https://doi.org/10.3390/md19020070

Academic Editor: Khaled A. Shaaban Received: 27 December 2020 Accepted: 26 January 2021 Published: 28 January 2021

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**Copyright:** © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

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as marine enzymes have demonstrated their useful for both process improvement and for the development of new process or products. Relevant types of lipases from marine organisms were identified and their novel features were discussed. They display, for example, salt tolerance, calcium independence and thermostable activities; they can also be stable in alkaline environment and were suggested to have antibiofilm action and higher catalytic efficiencies at temperatures lower than those from terrestrial microbial and/or mammal lipases [8]. However, few microalgal lipases and genes encoding lipases have been investigated and compared to bacterial, fungal, animal and plant lipases. In 2010, Demir and Tukel isolated and characterized for the first time a lipase from the photosynthetic cyanobacterium *Arthrospira. platensis* [9]. The lipase was a monomeric protein of 45 kDa with an isoelectric point of 5.9. It was specific for the 3-position in the ester bond. Godet et al. [10] isolated a new gene from the microalgae *Isochrysis galbana* encoding a 49 kDa lipase that shares similarities with fungal known lipase sequences. *Chlorella vulgaris* is a microalga belonging to the order of the *Chlorococcales*, which has a green color. It contains a significant number of intracellular proteins, carbohydrates, lipids, vitamin C, β-carotenes and B vitamins (B1, B2, B6 and B12), which is why it is commonly used for the preparation of food supplements. It is considered as raw materials for chemical compounds that have been affected by its primary and secondary metabolism, such as lipids, whose main application is for the generation of biodiesel [11]. This microalga has one of the highest lipid accumulating abilities in microalgae (50% of its DW), very high volumetric lipid productivity (VLP) of about 80 mg/L.day with a high growth rate in large-scale outdoor cultivation systems. Genetic manipulation technique for this microalga has already been established, showing grea<sup>t</sup> promise for improving its oleaginous phenotype by metabolic engineering [12]. Recently, its whole genome sequence was revealed by next-generation sequencing technologies, and the major metabolic pathways were identified [13]. Lipid metabolism has also been analyzed in multi-omics studies, including transcriptomics and proteomics to obtain the mechanistic insight of its lipid biosynthesis [14]. However, the TAG lipases have not been investigated yet. It will be of grea<sup>t</sup> importance to estimate the number and characteristics of its lipases, attracting knockdown targets for enhancement of lipid productivity. Here, a bioinformatic screening of a *C. vulgaris* genome was done to explore the presence of genes encoding putative lipases. The potential properties of the candidates are discussed on the basis of their three-dimensional (3D) model structures.
