The present invention relates to a method of producing a biologically active polypeptide having insulinotropic activity, the method comprising steps of: (a) transforming a genetically modified host cell that has protease gene knockout, with a polynucleotide vector encoding the polypeptide； and (b) growing the transformed host cell to produce the biologically active polypeptide； and a method of producing a biologically active polypeptide having an N-terminal recognition site His-Gly with insulinotropic activity, the method comprising steps of: (a) transforming a genetically modified Pichia pastoris that has protease gene STE13 knockout, with a polynucleotide vector encoding the polypeptide； and (b) growing the transformed Pichia pastoris to produce the biologically active polypeptide.
