Production of Laccase by Recombinant Yarrowia lipolytica from Molasses: Bioprocess Development Using Statistical Modeling and Increase Productivity in Shake-Flask and Bioreactor Cultures

Abstract

Laccases are used in numerous applications, from green degradation of various xenobiotic compounds, waste detoxification, textile dye bleaching, and delignification of lignocellulose materials to biofuel production. In this study, the recombinant Yarrowia lipolytica YL4 strain carrying the white-rot fungus Trametes versicolor laccase IIIb gene was used for laccase production from beet molasses as an agro-industrial residue. Response surface methodology was used to statistical optimization of the production of laccase by Y. lipolytica using an industrial medium containing molasses which allows a six times increase in laccase activity compared to primary medium contains glucose after 144 h. In bioreactor cultivation after 48 h, laccase production reached to 3.7- and 22.5-fold more than optimized and primary media in shake-flask cultures, respectively. Laccase productivity in bioreactor (0.0937 U/h) was higher than shake-flask culture (0.0084 U/h). The present study provides valuable information about statistical optimization of bioprocess development for cost-effective production of laccase and other heterologous proteins in Y. lipolytica from beet molasses as sole carbon source, thus allowing the valorization and decreasing environmental pollution of this agro-industrial waste.

Valuation of agro-industrial wastes as substrates for heterologous production of α-galactosidase

“…For the bioreactor culture, Q p was 0.158 U/L/h, notably higher than the other shake flask cultures and Y p /s was similar to the culture in shake flasks with YPW and also higher than the other cultures. The improved productivity in bioreactors is a common trait, generally attributed [4] to the fact that bioreactors are closed systems with a stricter control of aeration, agitation and temperature than other systems. Moreover, the mechanical agitation of bioreactors, compared to the orbital agitation of shake flasks, favors the continuous supply of oxygen to the cells and in consequence growth and bioproductions.…”

“…One outstanding bio-production is the microbial fermentation to ethanol [1, 2] or other biofuels [3] as an alternative to current threat of fossil fuels depletion and growing environmental concerns associated to their massive consumption. The production of heterologous proteins, including enzymes, using microbial cell factories is also remarkable [4]. …”

Biovalorization of vegetable oil refinery wastewater into value‐added compounds by Yarrowia lipolytica

“…The software recommended 30 experiments to perform on the basis of concentrations and levels of variables (Table ). After preparation of 30 trials, they were inoculated with seed culture (5% v/v) and incubated at 29 °C and 200 rpm for 48 h. Then, results of the experiments were inserted into the software for analysis and determine the best concentrations and levels of variables in order to obtain the maximum microbial oil by Y. lipolytica …”

“…A 200 mL seed was cultured in a 1000‐mL flask and kept at 200 rpm and 29 °C for 24 h. Then it was transferred to the bioreactor at an initial optical density (OD) at 600 nm of 0.1. The culture in the bioreactor was done with an airflow rate of 1 vvm at 500 rpm and 29 °C for 20 h. pH was adjusted to 6 by addition of KOH and H 3 PO 4 Mol L −1 …”

Pilot scale production of extracellular thermo‐alkali stable laccase from Pseudomonas sp. S2 using agro waste and its application in organophosphorous pesticides degradation

“…Agricultural waste is one of the alternative substrates that can be used for laccase production, also solving problems of disposal as well. In addition to this agrowaste also support the growth of microorganisms by giving essential nutrients, and work as an inducer for enzyme production in high titer …”