Studies of P450 enzymes of the CYP152 family capable of promoting the decarboxylation of fatty acids

Author(s)

Isabelle Taira Simões

• Document type: Master's Dissertation
• Press: Campinas, SP.
• Institution: Universidade Estadual de Campinas (UNICAMP). Faculdade de Ciências Farmacêuticas
• Defense date: 2024-07-23

Examining board members

Letícia Maria Zanphorlin Murakami; Danielle Biscaro Pedrolli; Priscila Oliveira de Giuseppe

• Advisor: Letícia Maria Zanphorlin Murakami; Wesley Cardoso Generoso

Abstract

The urgency to develop more sustainable alternatives to replace petroleum derivatives has grown considerably in recent years. This demand is driven by fluctuations in the price of crude oil, irreversible reduction in reserves, rapid increase in global energy demand, and concerns related to the environmental imbalance generated by the extraction and use of these substances. Thus, the in-depth study of biological routes capable of producing hydrocarbons, the main petroleum component, from vegetable raw materials becomes an interesting option. The discovery of P450 peroxygenase proteins gained notoriety due to their ability to catalyze the removal of oxygen from medium- to long-chain fatty acids, producing hydrocarbons. These enzymes, known as OleTs, catalyze the synthesis of alkenes with a double bond at carbon ?, from the free fatty acid decarboxylation reaction. However, little is known about these proteins and some challenges need to be overcome, such as obtaining enzymes with preferential decarboxylation activity over hydroxylation, since P450 peroxygenases can catalyze both reactions. In this sense, understanding the reaction mechanisms of P450 decarboxylases and the discovery of different enzymes with this activity is crucial for developing new technologies. The present dissertation therefore proposed the in-depth investigation of new putative P450 peroxygenases with possible decarboxylase functions and their applications in the in vitro and in vivo production of alkenes. The investigated enzymes were selected from in-silico analyses using sequence similarity networks (SSN). This analysis defined isofunctional clusters and used as niches to search for peroxygenases with decarboxylation activity. The data obtained from biochemical characterizations indicated that the enzymes studied have different specificities between them and others reported in the literature. As a highlight, the peroxygenase from Aeromicrobium yanjiei (OleTAer), from a cluster never explored, demonstrated great decarboxylase capacity, with conversion rates of different fatty acids that even surpass data already described in the literature. These results made it possible to use this OleT in the construction of strains of the oleaginous yeast Yarrowia lipolytica for the in vivo production of alkenes. Therefore, the present work demonstrated the potential of decarboxylase enzymes from the CYP152 family for the sustainable production (in vitro and in vivo) of hydrocarbons from fatty acids (AU)

• FAPESP's process: 21/14410-2 - Study of P450 enzymes of the CYP152 family able to promote the decarboxylation of fatty acids
• Grantee: Isabelle Taira Simões
• Support Opportunities: Scholarships in Brazil - Master