The invention discloses a method for enhancing the glucose oxidase secretion by transforming folding and secreting pathway of a protein, and belongs to the technical field of gene engineering. KAR2, CNE1, ERO1, SSA4, SSO2, SEC53, BMH2, HRD1, UBC1, GCN4 and other genes in Pichia pastoris are respectively cloned and linked into pGAPZ series derived plasmids of a Pichia pastoris expression vector bya gene recombination technology, the obtained plasmids and GOD was co-expressed and transferred into Pichia pastoris GS115 strains, the effect of the strain is investigated, modular combination optimization is carried out, and the strains are screened and identified to obtain a strain having a higher glucose oxidase secretion ability than the original strains. The enzyme activity of the glucose oxidase expressed by the strain in a 3 L fermentation tank is 1972.9 U / mL. The method lays a good foundation for the large-scale production of the glucose oxidase.