Process design and modeling for the production of triacylglycerols (TAGs) in Rhodococcus opacus PD630

Abstract

The oleaginous microorganism Rhodococcus opacus PD630 was used to study the characteristics and kinetics of the accumulation of triacylglycerols (TAGs) in cells. In this process, accumulation of TAG is stimulated when a carbon source is present in the medium in excess and the nitrogen source is limiting growth. Under controlled fermentation conditions the organism Rhodococcus opacus PD630 has been shown to grow to high cell density, producing high yields of TAGs (above 50% of cell dry weight) in a relatively short period of time. In this study, the reaction stoichiometry was established and the carbon balance for the process has been effectively closed, accounting for approximately 91% of the total carbon in the system. Several fed-batch strategies were explored at the IL benchtop bioreactor scale. Feeding both carbon and ammonium sulfate as the nitrogen source can sustain cell growth but was found to significantly obstruct the accumulation of TAGs. While these fed-batch strategies did not lead to titer improvements, they did highlight the significance of TAG degradation for growth. To aid in future process design strategy optimization an unstructured kinetic model was developed to describe the dynamics of the fermentation of Rhodococcus opacus PD630 and its triacylglycerol (TAG) production. The kinetic parameters for this model were either measured from experimental data or estimated by fitting the experimental data using least-squares non-linear regression. Global minimum of the sum of squared errors (SSE) between the model prediction and various experimental data sets was found by an iterative process of parameter space exploration. The minimum SSE obtained was 91.229. The proposed model is the first step towards understanding and optimizing the process of lipid production and accumulation in oleaginous organisms.