Production, purification and characterization of human GCSF protein in soluble form with co-expression

Human granulocyte colony stimulating factor (GCSF) is a hematopoietic growth factor and plays a crucial role in neutrophil production and differentiation. GCSF, which has a very important place in the clinic, is used to help the immune system up to the level of neutrophils required by the immune system, such as chemotherapy-induced neutropenia and AIDS treatment. Some foreign biomolecules, especially of human origin, such as GCSF, sometimes aggregate because of different factors during expression and creates inclusion bodies in E.coli expression system. Refolding process is often used to recover these very valuable molecules, but still significant amounts of protein remain unusable. Refolding processes are both costly, time consuming and not fully efficient. For these reasons, the use of chaperone proteins by co-expression method has been considered in this thesis study. In this context, using five chaperone plasmid systems (pG-KJE8, pGro7, pKJE7, pG-Tf2, pTf16) it has been tried to produce GCSF in soluble form. As a result of the experiments, it was found that pKJE7 plasmid is more effective in obtaining GCSF in soluble form. GCSF protein co-expressed with this system was obtained in high purity by affinity chromatography and its proliferative activity on human umbilical cord endothelial cells (HUVEC) was demonstrated. As a result, GCSF, which previously aggregated as an inclusion body in the E. coli expression system, was correctly folded by co-expression with chaperone proteins and was obtained as functional and pure.