Aim To investigate the high level expression of human phospholipid transfer protein in the yeast pichia pastoris. Methods Total RNA was prepared from Chinese fetal liver tissue, cDNA fragment encoding human phospholipid transfer protein was amplified by reverse transcriptase polymerase chain reaction using specific primers, and then was integrated into the chromosomes of pichia pastoris GS115 via homologous recombination. Recombinant human phospholipid transfer protein was expressed under the control of the promoter of the alcohol oxidase gene(AOX1) and was detected by SDS-PAGE. Results Five recombinant colony among 20 colonies grown poorly on MM plates was selected which highly expressed human phospholipid transfer protein. The molecular weight of recombinant phospholipid transfer protein was estimated about 75 kDa analyzed by SDS-PAGE. Conclusions Human phospholipid transfer protein can be high level expressed in the yeast pichia pastoris and the secreted recombinant human phospholipid transfer protein was in the form of the glycosylated monomer.