Integration of heterologous 4-hydroxybenzoic acid transport proteins in Rhodobacter sphaeroides for enhancement of coenzyme Q10 production

Abstract

The highly redox-active compound, Coenzyme Q₁₀ (CoQ₁₀), has attracted increasing interest due to its clinical benefits and important applications in the pharmaceutical, food and health industries. Although the metabolic pathways involved in CoQ₁₀ biosynthesis are known, very few reports are available concerning the development of engineering strategies to enhance CoQ₁₀ production in the natural producer Rhodobacter sphaeroides. In this work, three membrane transport proteins, AcPcaK, KpPcaK, and CgPcaK from different organisms (Acinetobacter calcoaceticus, Klebsiella pneumoniae and Corynebacterium glutamicum, respectively) were individually heterologously expressed in R. sphaeroides GY-2 to enhance the uptake of extracellular 4-hydroxybenzoic acid (4HBA), an important intermediate of CoQ₁₀ synthesis. Each of the PcaK open reading frames (ORFs) was inserted behind an RFP gene to weaken their expression in order to avoid inhibition of cell growth. The transport efficiency and consumption of 4HBA, as well as CoQ₁₀ productivity of the resulting recombinant R. sphaeroides GY-2 strains has been studied. The CoQ₁₀ productivity of RS-CgPcaK and RS-KpPcaK was effectively improved by the addition of external 4HBA, reaching maximum values of 17.45 and 18.06 mg g⁻¹ DCW (18.05% and 20.82% higher than R. sphaeroides GY-2), respectively. The strategy used in this study proved effective for enhancing the biotechnological production of CoQ₁₀ and thus holds promise for further improvements, including industrial applications.