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

Aggregation of misfolded proteins can reduce the yield in recombinant protein 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 properly the specific concentration of a human protein produced in E. coli and its partitioning into soluble and insoluble cell fractions. Accurate estimation of the protein-specific parameters required informative experiments, which were designed using the Fisher information matrix. The model was employed to calculate the influence of the specific glucose uptake rate in high-cell-density cultivation of E. coli on accumulation and aggregation of the recombinant protein. Despite its simplicity, the model was flexible and unbiased concerning unidentified mechanisms. Assuming an exponentially decreasing production rate, the irreversible aggregation step was found to follow first order kinetics, while assuming a constant production rate with simultaneous degradation, the model predicted transient aggregation only. Implications for strain and process development are discussed. Copyright 2001 John Wiley & Sons, Inc.