Codon bias was one of the important parameter which influence heterogenous gene expression, optimizing codon sequence could improve expression level of heterogenous gene. In the preview study, wildtype E2 gene was expressed poorly in Pichia pastoris, in order to improve the expression level of E2 gene in Pichia pastoris, the low usage codons of E2 gene were mutated into high usage codons in Pichia pastoris by directed-mutagenesis based on PCR. The result showed that, compared with the results reported in preview study, the expression level of E2 gene in Pichia pastoris was improved observably by substituting 24 low usage codons of E2 gene for the high usage synonymous codons. It suggested the stragety to improve the expression of E2 gene in Pichia pastoris by codon optimization was successful.