Production of Leptin inEscherichia coli:A Comparison of Methods

A procedure is described for gram-scale refolding ofEscherichia coli-derived human leptin inclusion bodies. Refolding was achieved by gradually reducing denaturant using a diafiltration method. Refolded leptin is characterized byin vivomodulation of food intake, reduction in body weight, and lowering of insulin and glucose levels inob/obmice. In addition, refolded leptin is characterized by radioimmunoassay (RIA) and activation of the leptin receptor in a cell-based assay. For comparison we also refolded leptin by a simple dilution method and produced periplasmic derived leptin, which did not requireex vivofolding. Leptin produced by these three methods and leptin obtained from commercial sources were compared using the RIA and the cell-based assay and appeared to be of comparable quality and potency.