Studies On Soluble Expression Of Recombinant Proteins In Escherichia Coli

Large amount of pure, soluble and functional proteins are of high demand in the academic, industrial and pharmaceutical fields. The proteins from natural resources are however unable to meet these requirements, due to the limited contents and high expense for isolation and purification. With the development of DNA recombinant technology and protein engineering, the recombinant proteins conferring the physical, chemical and biological features similar to the natural ones, can be obtained by large-scale preparations. So far, several expression systems for recombinant protein production are available, by using different hosts including the prokaryotic Escherichia coli (E. coli) and the eukaryotic yeast, insect and mammalian cells. Therein, the E. coli expression system has been prevalently used, especially for high throughput expression of the recombinant proteins relating to the current vigorous studies in the proteomics. This system has many advantages including well-known genetic background, simple culturing conditions, fast growth, low cost and large-scale production, and incomparable availability of numerous potent expression vectors, strains and purification systems. Nonetheless, the E. coli expression system has been practically hindered to a large extent by its inherent defect that the expressed recombinant proteins frequently formed insoluble and non-active aggregates termed inclusion bodies, requiring a laborious and inefficient process for denaturalization and refolding. To solve this problem, a dozen of manners to improve the protein solubility have been raised, with the increased understanding of the protein folding mechanism in E. coli. Among them, the fusion expression strategy is a better choice. In this study, a set of distinct protein fusion partners were comparatively evaluated to promote the soluble expression of two target proteins in E. coli including the bovine enterokinase largely prone to aggregation and the green fluorescent protein with moderate active solubility. Within protein attributes that are putatively involved in protein solubility, the protein acidity is of particular concern. Our results explicitly indicated the protein fusion partners with a stronger acidity remarkably exhibited a higher capacity to enhance the solubility of the target proteins. Among them, MsyB, an E. coli acidic protein that suppresses the mutants lacking function of protein export, was revealed as an excellent protein fusion partner with the distinguished features including highpotential to enhance protein solubility, efficient expression, relatively small size and the origin of E. coli itself. In principle, our results confirmed the modified solubility model of Wilkinson-Harrison and especially deepened understanding its essence. Meanwhile, the roles of other parameters such as protein hydrophilicity in solubility enhancement were discussed, a guideline to design or search an optimum protein solubility enhancer was also proposed.