Lipoxygenase : a game-changing enzyme

Many challenges lie ahead in using LOXs as tools in industrial oleochemistry. One of these challenges is the supply of PUFAs. Although we are moving towards a biobased economy where second and third generation biomass is taking a leading role, it is still faster and cheaper to use first generation biomass. Industrialization of microbial oils is a good alternative to supply the demand of PUFAs. Another challenge is the production of heterologous LOX in sufficient quantities. Since the last decade this problem is being tackled and more research is being done in heterologous expression of LOXs. The LOX with the highest potential so far is the secreted <em>Pseudomonas aeruginosa</em> LOX produced in <em>Escherichia coli</em>. During this thesis research different <em>lox</em> genes were tried for heterologous production of LOX using different <em>Aspergillus niger</em> and <em>Aspergillus nidulans</em> strains as expression hosts. These LOXs were identified as discussed in <strong>Chapter 3</strong> and <strong>Chapter 6</strong>. Unfortunately, heterologous production in sufficient quantities was unsuccessful using these expression hosts as discussed in <strong>Chapter 5 </strong>and <strong>Chapter 6</strong>. Since production of <em>Gaeumannomyces graminis</em> LOX was successful in <em>Trichoderma reesei</em>, as discussed in <strong>Chapter 4</strong>, the production of polymers used for bioplastics could be demonstrated in this ERA-NOEL project anyway. Therefore this thesis shifted its focus on resolving the question of the difficulties in the heterologous expression of LOX in different <em>Aspergillus </em>species. <strong>Chapter 5</strong> is the result of a systematic approach to analyze different aspects of <em>G. graminis </em>LOX expression in <em>A. nidulans</em>. <strong>Chapter 2</strong> shows that heterologous expression of extracellular fungal LOX can be performed using <em>T. reesei</em> and <em>Pichia pastoris</em> as production hosts, and <em>E. coli</em> can be used for the production of intracellular LOXs of plant, mammal, bacterial, and fungal origin. As shown in <strong>Chapter 2</strong>, <em>E. coli </em>is not very efficient in the production of heterologous LOX due to the formation of inclusion bodies and low induction temperature necessary for production. The use of <em>Aspergillus oryzae</em> can be exploited further in the heterologous production of LOXs. Due to the choice of using <em>A. niger</em> and <em>A. nidulans</em> as expression hosts, this expression host was not exploited for its potential. The last challenge is to synthetically engineer LOX to broaden its use in industry. In this way more building blocks for chemicals can be synthetically produced and more products based on LOX origin can be made. Therefore, LOX can be a world-wide game-changing enzyme in a biobased economy as its use can decrease the demand for petroleum-based products.