Purification Of GGH Fusion Protein And Preliminary Evaluation Of Pharmacodynamical

Glucagon-like peptide-1 (GLP-1) is 30-residual peptide hormone secreted by intestinal L-cells. It can not only stimulate pancreatic beta cells proliferation but also inhibit pancreatic beta cells apoptosis; including inhibition of both gastric empting and food intake. Its half-life in circulation is very short and provided mainly by fast renal clearance due to its low molecular weight and proteolysis susceptibility, resulting in low clinical utility. In our lab, A GLP-1 fusion protein named GGH comprising double tandem GLP-1 and human serum albumin(HSA) was constructed to prolong its half-life. In this study, the purification of fusion protein GGH was optimized, and pharmacodynamic evaluation of the protein was performed. The main conclusions are as follows:The GGH was purified from fermentation broth by centrifugation, ultrafiltration concen -tration, affinity absorption chromatography, gel filtration and ion exchange chromatography. The final product was conformed as one single band by SDS-PAGE and the purity was identified as 98.48%. The total recovery yield could reach up to 28.9%. Endotoxin of GGH was removed and to a concentration of 10 EU/mg.Method to observe minimum effective dose, maximum tolerance dose, dose-response relationship adiministered as a single intravenous injection, dose-response and time-response relationship as a single subcutaneous injection on oral glucose tolerance test (OGTT). Fusion protein of GGH at the dose of 1.0~6.0 mg/Kg, administered as a single intravenous or subcutaneous, lowered plasma glucose in the linear relationship; the best effect after 8 h subcutaneous injection.Fusion protein of GGH evaluate pharmaceutical on the model ofβcell damaged and on the normal mice of gastric emptyting. In immunohistochemical results, compared with model group, GGH at the dose of 2.0, 6.0, 18.0 mg/Kg could increase the level of insulin and the contents of muscle and liver starch; decreased the diet and water of model mice;stimulated pancreatic beta cells proliferation and inhibited pancreatic beta cells apoptosis. At the dose of 2.0, 4.0, 6.0 mg/Kg, GGH could significant inhibit both of gastric empting and food intake.