Genomic and Molecular Characterization of Botrytis cinerea Lipase for Investigating its Biosynthetic Potential

Abstract

Lipases are known to degrade the lipids in the form of fatty acids and microbial lipases have shown more potential toward the enhanced enzyme yield. However, there is paucity of data on fungal lipases that could be utilized for commercial applications. Current study was aimed to explore the molecular and genomic insights into a phytopathogenic fungus, B. cinerea for its lipase biosynthetic potential. Approximately 161 plant based fatty acid molecules were docked with B.cinerea lipase to be evaluated in accordance with their resultant binding affinity values for biofuel synthesis. Among the selected triglycerides, docosahexaenoic acid, dicranin and hexadeca7,10,13-trienoic acid were subjected to molecular dynamic simulations for 20 ns. Three selected candidates were found potent for biofuel production. Mycoviruses are known to cause different impact on fungal host including hypervirulence, hypovirulence as well as cryptic response. We hypothesized that the mycoviruses may have regulatory role in the lipase production in vitro. Hence to further investigate, we selected virus infected Kst5C, Kst14A, Kst32B, Kst33A, Kst58 and Kst31C) and virus free(Kst51) botrytis isolates showing variable lipase production. All isolates were characterized on morphological, biological and at molecular level. The hypovirulent strain behaves distinctly than virus free and hypervirulent strain in terms of growth patterns, pathogenicity, and morphologic appearance thus showing differential role in their respective host. GCMS data of all B. cinerea strains were found rich in metabolic profiles with various bioactive compounds in the form of organic acids, polyketides, alcohols, fatty acids and many other with a relative difference. B. cinerea lipase optimal growth was observed on 25°C temperature, 6.5 pH, olive oil-based fermentation medium enriched with glucose and 3.5ml inoculum for 15 days. The hypervirulent strain Kst58 showed maximum mycelial growth, higher pathogenicity, higher alcoholic contents as well as higher lipase activity (156.82 U/ml), followed by virus free, Kst51 (148.42U/ml) and Kst31C (139.41 U/ml). Mitochondria provide energy for lipase breakdown which is further used in production of energy rich ATP molecules thus helping cell metabolism inside mitochondria. We evaluated differences in four mitogenomes throughout the genomic characterization process, revealing that the isolates' size range of 55–77 kbp may account for their varying lipase production. And the isolate Kst14A with 77kbp genome was found to produce more lipase. Hence, B. cinerea mycovirus infected isolate Kst58 and Kst14A can be utilized as a potential candidate for exploration of bioactive compounds leading towards higher production of extracellular enzyme lipases to be used for commercial purpose.