| Institution: | aBiomolecular Science, University of Science and Technology, Daejeon 305-350, Republic of Korea;bAdvanced Analysis Center, Korea Institute of Science and Technology, Seoul 136-791, Republic of Korea;cDepartment of Pharmacy, CHA University, Gyeonggi-do 487-801, Republic of Korea;dBiomedical Center, Korea Institute of Science and Technology, Seoul 36-791, Republic of Korea;eResearch Institute of Pharmaceutical Sciences, College of Pharmacy, Seoul National University, Seoul 151-742, Republic of Korea |
| Abstract: | STAM1 and Hrs are the components of ESCRT-0 complex for lysosomal degradation of membrane proteins is composed of STAM1 Hrs and has multiple ubiquitin binding domains. Here, the solution structure of STAM1 UIM, one of the ubiquitin binding motif, was determined by NMR spectroscopy. The structure of UIM adopts an α-helix with amphipathic nature. The central hydrophobic residues in UIM provides the binding surface for ubiquitin binding and are flanked with positively and negatively charged residues on both sides. The docking model of STAM1 UIM-ubiquitin complex is suggested. In NMR and ITC experiments with the specifically designed mutant proteins, we investigated the ubiquitin interaction of tandem ubiquitin binding domains from STAM1. The ubiquitin binding affinity of the VHS domain and UIM in STAM1 was 52.4 and 94.9 μM, and 1.5 and 2.2 fold increased, respectively, than the value obtained from the isolated domain or peptide. The binding affinities here would be more physiologically relevant and provide more precise understanding in ESCRT pathway of lysosomal degradation. |
