To identify the interacting proteins of transactivation domain of forkhead box protein M1 which is located on the 688th to the 748th amino acid sequence of C-terminal end，the cDNA of FOXM1 was used as a template, the transcription activation domain sequence was amplified by polymerase chain reaction and cloned into the prokaryotic expression vector pGEX-4T2. The recombinant protein GST-FOXM1688-748 fused with glutathione S- transferase tag was obtained by Escherichia coli prokaryotic expression. The interacting proteins were identified by GST-pulldown assay and mass spectrometry. The pGEX-4T2-FOXM1688-748 prokaryotic expression plasmid was constructed successfully and the FOXM1688-748 recombination protein was obtained. According to the mass spectrometry results, the interacting proteins were analyzed and classified to show that some of them activated the transcriptional activity of FOXM1, such as RPN2, MISP, and MCM7 proteins, and some of them regulated the stability of FOXM1 protein, such as USP9Y, CUL4A, HSPB1, and BAG2 proteins. The transcription activation domain of FOXM1 protein interacts with many proteins participating in different pathways, indicating that this domain has an important molecular biological role, which can expect to provide experimental basis for clinical drug development targeting FOXM1.