Oxidation of propionate in Pseudomonas sp. LFM046: Relevance to the synthesis of polyhydroxyalkanoates containing odd-chain length monomers and 2-methylisocitrate

Pedro Henrique Santos-Oliveira; Nathália Fernandes Gonçalves Machado; Rafael David de Oliveira; Eric André Velasco-Yépez; Sonia Regina da Silva; Rafael Costa Santos Rocha; Lars Mathias Blank; Luiziana Ferreira da Silva; Galo Antonio Carrillo Le Roux; José Gregório Cabrera Gomez

doi:10.1016/j.biortech.2023.129944

Oxidation of propionate in Pseudomonas sp. LFM046: Relevance to the synthesis of polyhydroxyalkanoates containing odd-chain length monomers and 2-methylisocitrate

Bioresour Technol. 2023 Oct 28:129944. doi: 10.1016/j.biortech.2023.129944. Online ahead of print.

Affiliations

¹ Departamento de Microbiologia, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo, Brazil; iAMB - Institute of Applied Microbiology, ABBt - Aachen Biology and Biotechnology,RWTH Aachen University, Worringerweg 1, 52074 Aachen, Germany.
² Departamento de Microbiologia, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo, Brazil.
³ Department of Chemical Engineering, Norwegian University of Science and Technology (NTNU), Trondheim, Norway.
⁴ Global Center Research Innovation and BioSolutions, Rovensa Next, Brazil.
⁵ iAMB - Institute of Applied Microbiology, ABBt - Aachen Biology and Biotechnology,RWTH Aachen University, Worringerweg 1, 52074 Aachen, Germany.
⁶ Departamento de Engenharia Química, Escola Politécnica, Universidade de São Paulo, São Paulo, Brazil.
⁷ Departamento de Microbiologia, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo, Brazil. Electronic address: [email protected].

PMID: 39492536
DOI: 10.1016/j.biortech.2023.129944

Abstract

Pseudomonas sp. LFM046 produces polyhydroxyalkanoates of medium-chain length. When carbohydrates are used, only monomers of even-length chains (3HA_even) are generated. Propionate, when used as a co-substrate, enabled the synthesis of odd-chain length monomers, albeit with poor yields (Y_3HAodd/prop.) of approx. 10 %_. A mini-Tn5 mutant (LFM693) was generated by interrupting the prpB gene, which encodes the 2-methylisocitrate lyase responsible for one step in propionate oxidation via 2-methylcitrate cycle. At low propionate concentrations of 0.1-0.3 g.L^-¹, LFM693 showed higher Y_3HAodd/prop. In contrast, the propionyl incorporation into the polymer was less efficient at high propionate concentrations. The excretion of 2-methylisocitrate partially explained this reduced efficiency. However, mass balances and ¹³C isotopomer analysis revealed significant propionate oxidation in LFM693 of 21 % and 46 % for propionate concentrations of 0.5 and 1.0 g.L^-¹, respectively. Experiments using C1-labeled (¹³C) propionate revealed an alternative propionate oxidation pathway (α-oxidation, β-oxidation, or 3-methylmalate) whose genes are yet to be revealed.

Keywords: 2-methylcitrate cycle; 2-methylisocitrate; Metabolic engineering; Polyhydroxyalkanoates; Propionate oxidation.