Identification of a novel ubiquitin binding site of STAM1 VHS domain by NMR spectroscopy

Abstract 

Interaction between the signal-transducing adapter molecule 1 (STAM1) Vps27/Hrs/Stam (VHS) domain and ubiquitin was investigated by nuclear magnetic resonance (NMR) spectroscopy. NMR evidence showed that the structure of STAM1 VHS domain resembles that of other VHS domains, especially the homologous domain of STAM2. We found that the VHS domain binds to ubiquitin via its hydrophobic patch consisting of N-terminus of helix 2 and C-terminus of helix 4 in which Trp26 on helix 2 plays a pivotal role in the binding. The binding between VHS and ubiquitin seems to be very similar to that between ubiquitin associated domain (UBA) and ubiquitin, however, the direction of α-helices involved in the ubiquitin binding is opposite. Here, we propose a novel ubiquitin binding site and the manner of ubiquitin recognition of the STAM1 VHS domain.

Structured summary

MINT-6804185:

STAM1 (uniprotkb:Q92783) binds (MI:0407) to ubiquitin (uniprotkb:P62988) by nuclear magnetic resonance (MI:0077)

Abbreviations: STAM, signal-transducing adapter molecule, VHS, Vps27/Hrs/Stam, NMR, nuclear magnetic resonance, UBA, ubiquitin associated domain, ESCRT, endosomal sorting complex required for transport, UIM, ubiquitin interacting motif, GAT, GGAs and TOM, MVB, multivesicular body, SH3, Src homology domain 3, GGA, golgi-localizing/γ-adaptin ear homology domain/ADP-ribosylation factor-binding protein, Hrs, hepatocyte growth factor-regulated substrate, Tom, target of Myb, Srcasm, Src-activating signaling molecule, TGN, transgolgi network, PDB, protein data bank, GST, glutathione S-transferase, IPTG, isopropyl β-d-1-thiogalactopyranoside, DTT, 1,4-dithiothreitol, PMSF, phenylmethanesulphonylfluoride, HSQC, heteronuclear single quantum correlation spectroscopy, DSS, 2,2-dimethyl-2-silapentane-5-sulfonic acid, CSI, chemical shift index, NOE, nuclear overhauser enhancement, K_d, equilibrium dissociation constant, CUE, coupling of ubiquitin conjugation to endoplasmic reticulum degradation, MIU, motif interacting with ubiquitin

Keywords: STAM1 VHS domain, Ubiquitin recognition, NMR spectroscopy, Chemical shift perturbation, Protein–protein interaction