Kinetic model of in vivo folding and inclusion body formation in recombinant Escherichia coli

Aggregation of misfolded proteins can reduce the yield in recombinant prote in production. The underlying complex processes are additionally influenced by cellular physiology. Nevertheless, a lumped-parameter model of kinetic competition between folding and aggregation was sufficient to track properl y the specific concentration of a human protein produced in E. coli and its partitioning into soluble and insoluble cell fractions. Accurate estimatio n of the protein-specific parameters required informative experiments, whic h were designed using the Fisher information matrix. The model was employed to calculate the influence of the specific glucose uptake rate in high-cel l-density cultivation of E. coli on accumulation and aggregation of the rec ombinant protein. Despite its simplicity, the model was flexible and unbias ed concerning unidentified mechanisms. Assuming an exponentially decreasing production rate, the irreversible aggregation step was found to follow fir st-order kinetics, while assuming a constant production rate with simultane ous degradation, the model predicted transient aggregation only. Implicatio ns for strain and process development are discussed. (C) 2001 John Wiley & Sons, Inc.