Investigation of Nitrate Respiration in Cupriavidus necator for Application in Life Support System

Abstract

Cupriavidus necator is a well-studied microorganism with potential application in bioregenerative life support systems for single-cell protein and bioplastic production. Most studies have been carried out in autotrophy or heterotrophy, requiring O2 as the final electron acceptor. In the context of inhabited missions, access to O2 will primarily be limited to the crew. In this study, we investigated the capacity of C. necator to carry out nitrate respiration as a strategy to limit oxygen supply to the cultures by providing nitrate from another compartment of the Bioregenerative Life Support System (BLSS). Batch bioreactor experiments were carried out to determine the best conditions for nitrate utilization in terms of pH and aeration. Continuous cultures were then performed under two carbon sources (glucose vs. acetic acid) and two substrate limitations (nitrate vs. carbon). The optimal conditions were found to be pH 7.5 under anaerobiosis. They were applied in chemostats, where three steady-states were obtained at a low dilution rate. In all cases, the biomass consisted of a mixture of protein (from 29 ± 1% Cell Dry Weight (CDW) to 39 ± 2% CDW) and polyhydroxybutyrate (from 45 ± 2% CDW to 57 ± 3% CDW), which was found to be a key component for nitrate respiration metabolism. Microaerobic conditions were also tested in batch culture, reporting for the first time aerobic nitrate respiration in C. necator. Under these conditions, growth parameters improved during the nitrate phase; however, the specific growth rate during the nitrite phase was lower than that observed under strictly anaerobic conditions.