Human cytoplasmic alpha-fetoprotein (AFP) has been classified as a member of the albuminoid gene family.The protein sequence of AFP has significant homology to that of human serum albumin (HAS), but they have vastly different functions.The AFP functions as a regulator in the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway, but HSA plays a key role as a transport protein.To probe their molecular mechanisms, we have applied co-localization, co-immunoprecipitation (co-IP) and molecular docking approaches to analyze the differences between AFP and HAS.The data from co-localization and co-IP displayed a strong interaction between AFP and PTEN (phosphatase and tensin homolog), demonstrating AFP did bind to PTEN, but HAS did not.The molecular docking study further showed that the AFP could contact domains Ⅰ and Ⅲ of PTEN.In silicon substitutions of AFP binding site residues at position 490M/K and 105L/R in responding to residues K460 and R105 in HAS resulted in steric clashes with PTEN residues R150 and K46 respectively.These steric clashes may explain the reason why HAS cannot bind to PTEN.Ultimately, the experimental results and the molecular modeling data from the interactions of AFP and HAS with PTEN will help us to identify targets for designing drugs and vaccines against human hepatocellular carcinoma.