Peroxygenase-Catalyzed Selective Oxyfunctionalization to Produce Tetralols

Biocatalysis, in which enzymes are used as catalysts for chemical reactions, is emerging as a greener alternative to other types of catalysis that utilize toxic organic or org...ganometallic compounds. Enzymes are not only greener catalysts, but also highly chemo-, region-, and stereoselective. The main purpose of the research explained in this proposal is to utilize a peroxygenase, which has been extensively utilized in the laboratory of Prof. Frank Hollmann, the host faculty at Delft University of Technology, for selective oxidation in the last few years. Peroxygenases represent an important class of enzymes that are comparable to P450 monooxygenases, with the advantage that peroxygenases do not require the use of a cofactor. Herein, we propose to extend the application of a peroxygenase from Agrocybe aegerita (AaeUPO) in oxidation of selected tetralols to produce the corresponding tetralones, a transformation that has been reported to be impossible by alcohol dehydrogenases by several groups. The ability to oxidize these substrates with AaeUPO will enable the incorporation of this AaeUPO-catalyzed oxidation reaction with an alcohol dehydrogenase-catalyzed reduction in one pot, either sequentially or simultaneously, to accomplish a bienzymatic deracemization of racemic tetralols to their enantiopure form. The research group of Prof. Hollmann is a leading group in biocatalysis. The PI will spend two months of the summer of 2023 in this laboratory trying to accomplish this AaeUPO-catalyzed oxidation task, and learn few tricks related to the used AaeUPO, which utilizes hydrogen peroxide as a sacrificial substrate, in chemical reactions. The control of the amount of hydrogen peroxide that is generated in situ during the reaction is crucial to control the selectivity of peroxygenases in general, and AaeUPO in particular. Once the oxidation of tetralols is accomplished, this peroxygenase-based oxidation will be evaluated in terms of stereospecificity. If this reaction is proven stereospecific (i.e., depletes one enantiomer of a tetralol to the corresponding ketone leaving the other one untouched), it will be incorporated with an alcohol dehydrogenase-catalyzed stereoselective reduction of the resultant ketone. The selection of the ADH will be such as the stereopreference of this ADH meets the requirement of accumulation of the unreacted alcohol. This approach is known as deracemization via stereoinversion. If the oxidation using AaeUPO is shown to be non-stereospecific, a cyclic deracemization approach will then be used. The ability of AaeUPO to oxidized non-activated C-H bonds represents a very important approach for oxyfunctionalization of hydrocarbons. This is a long-term target objective for starting this collaborative work with the host faculty. This might contribute to a major objective for the IRC for Refining and Advanced Chemicals, where besides Chemistry Department Dr. Musa is currently affiliated to, which involve conversion of oil to chemicals. This option of oxyfunctionalization of non-activated C-H bonds is kept also as a plan B during this summer visit in case the intended selective oxidation of tetralols to their corresponding tetralones is shown impossible. Prof. Hollmann and the PI wrote a comprehensive review on deracemization approaches in 2019, and thus both are experts in the field of biocatalysis. Availability of fund for this summer program will allow the PI to get the proper training to handle a new enzyme. It will also enrich the laboratory of the PI at KFUPM with one more important enzyme with unique function. The host mentioned that he is also willing to share the plasmid of this enzyme anytime with the PI, which will guarantee the continuation of this collaboration and the advancement in the biocatalysis laboratory that is established in the Chemistry Department by the PI. This will contribute to assigning excellent projects to graduate students at the Chemistry Department of KFUPM that will guarantee proper training in the hot areas of biotechnology