1^st Speaker : Bekir Engin Eser

Title: Expanding the Toolbox of Fatty Acid Hydratases: A Thermostable Hydratase from Marinitoga piezophila with a Low Temperature Optimum and a Unique Regioselectivity

Abstract: Fatty Acid Hydratases (FAHs) catalyze the addition of water to unsaturated fatty acids to generate hydroxy fatty acids (HFAs) as products. Since HFAs have diverse application areas from materials and cosmetics industries and possess beneficial bioactivities, their benign enzymatic synthesis from abundant oils has attracted a lot of attention in the recent decade. One common challenge with biocatalytic conversions, including FAHs, is the stability of enzymes towards process conditions. Thus, we looked at Nature to find thermostable FAHs and characterized FAH ortholog from the thermostable and piezophilic organism Marinitoga piezophile. As expected, MpFAHY showed high thermostability, retaining over 90 % of its activity even after 30-min incubation at 70 °C. However, interestingly, the enzyme showed the highest activity at a much lower assay temperature of 20 °C, with sharp decreases above and below this temperature. This might indicate a physiological function of the enzyme, e.g. being part of a cold adaptation mechanism of the organism, which normally lives at 45-70 °C. Moreover, the purified enzyme requires NaCl to be active, consistent with the living habitat of its source organism. Another interesting property of the enzyme was its unique regioselectivity. MpFAHY was able to produce a mixture of 10-OH and 13-OH products from linoleic acid, with 13-OH being the preferred product, which is not a common property of wild-type FAHs.

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2^nd Speaker: Frederik Vig Benfeldt

Title: Multi-enzyme/whole cell catalytic production of short-medium chain diols and diacids 

Abstract: The rising issue of plastic pollution and the limited motivation for mechanical recycling urges the academia and industry to develop the technologies that enable close- or open- loop recycling and upcycling. To address this, the ACTPAC project seeks to develop a practical method to transform chemically inert C-C backboned plastic waste, specifically polyethylene (PE), into high-value monomers and biochemicals.One of the pivotal challenges in this transformation is the biotransformation of alkanes into α,ω-alkandiols and diacids, particularly due to the difficulty in directing C-H oxy-functionalization at the least reactive terminal positions. The ACTPAC project aims to utilize the unique capabilities of Cytochrome P450 (CYP450) enzymes to overcome this challenge. Our strategy involves screening and characterization of promising CYP153A orthologs capable of hydroxylating and oxidizing medium length alkanes into α, ω-diols and diacids. Engineering of selected CYP153A enzymes to fine-tune their specificity and activity towards targeted substrates through computational methods and machine learning algorithms. Lastly, scale-up is aimed to be conducted in large-scale bioreactors, with a focus on enhancing efficiency, cofactor regeneration, and fine-tuning reaction conditions to maximize yields.