Smad ubiquitination regulatory factor 1 (Smurf1) is an HECT type E3 ligase and regulating a lot of life activity, such as nervous development, cell polarization, bone remodeling and tumor formation, etc. At present, it is in-deep that our knowledge about Smurf1, but people did not particularly distinguish its isoform Smurf1 L and Smurf1 S (the former have 26 amino acid more than the latter) in previous study, and most of related work tend to Smurf1 S. So this article provided some information about the function of Smurf1 L and Smurf1 S. By means of RT-PCR, immunofluorescence assay and Western blot, we studied the difference of Smurf1 L and Smurf1 S in some aspects, including tissue expression, subcellular localization and substrate degradation. The result suggested that: on the one hand, not only the tissue distribution of Smurf1 S is more widespread than Smurf1 L, but also its expression level is higher than Smurf1 L; on the other hand, their subcellular localization have also obvious distinction. Smurf1 L exist at spindles in mitotic phase, but Smurf1 S perhaps exist in cytoplasm. In addition, the degradation ability to substrate of Smurf1 S is stronger than Smurf1 L, and have dose-dependent. In short, these achievement have important significance for more precise research to Smurf1.