Ubiquitin carboxyl-terminal hydrolase isozyme L3 (UCHL3) is a member of the ubiquitin C-terminal hydrolase (UCH) subfamily of deubiquitinating enzymes (DUBs) which regulate the ubiquitin-proteasome system by cleaving ubiquitin moieties from target proteins or ubiquitin precursors. UCHL3 exhibits diverse physiological functions including cell cycle regulation, the DNA damage response, and chromosome biorientation and segregation. Abnormal UCHL3 activity has been associated with the pathogenesis of various diseases, including breast, lung, and pancreatic cancers, yet the detailed molecular mechanisms underlying these functions remain poorly understood. Previous studies have suggested that the active sites of UCHL3 and other homologous UCHs exhibit substantial dynamics particularly in their enzymatic active site, which may be essential for ubiquitin recognition and enzymatic activity. To provide a structural basis for elucidating the dynamic behavior and functional mechanism of human UCHL3, here we report extensive NMR backbone and side-chain 1H, 13C, and 15N resonance assignments of the recombinant full-length human UCHL3 in solution.
The structural βγ -crystallin from the tunicate Ciona intestinalis serves as an evolutionary waypoint between ancestral metal cation-binding crystallins and the βγ crystallins of the vertebrate eye lens. Ci- βγ maintains the metal cation-binding sites of ancestral crystallins while also sharing common structural motifs with vertebrate lens crystallins. Due to its structure and strong interaction with Ca ^2+ , Ci- βγ is more thermally and chemically stable than vertebrate lens crystallins, making it an ideal starting point for understanding βγ crystallins’ evolution for molecular stability. In this study, we present the side-chain and updated backbone resonance assignments of Ci- βγ with TALOS-N predictions on the H _N , H _α , C _α , C _β and N chemical shifts. These assignments provide a foundation for future studies of metal binding, dynamics, and stability of proteins in the βγ -crystallin family.
The cellular export of large-sized proteins, such as collagen or fibrillin, necessitates a specialized transport pathway tailored towards their size. For collagen and possibly also other bulky proteins, the Transport ANd Golgi Organization 1 (TANGO1) protein plays a substantial role in the transfer from the Endoplasmic Reticulum (ER) to the Golgi apparatus. TANGO1 and related proteins organize the ER exit sites and generate large export structures within the cytosol while anchored to the ER membrane. In contrast to other proteins involved, TANGO1 features a folded domain in the ER lumen, which seems to be central for detecting cargos. This cargo-recognition domain has been shown to bind the collagen-specific chaperone HSP47 (Ishikawa et al. 2016) as well as type IV collagen directly (Arnolds and Stoll 2023). However, the exact molecular mechanism of these interactions remains unknown. The resonance assignments presented here lay the foundation for the experiments studying the binding process on a molecular level and is designed to facilitate investigations into how TANGO1 detects and selects the cargos to be exported.
The BRCA1-associated RING domain protein 1 (BARD1) is the obligate binding partner of the tumor suppressor breast cancer type 1 susceptibility protein (BRCA1) and plays a critical role in maintaining genome integrity. BARD1 contains structured N- and C-terminal domains that mediate heterodimerization with BRCA1, recognition of chromatin marks, and DNA repair functions. Approximately 40% of BARD1 is intrinsically disordered, particularly in the central region of the protein. This intrinsically disordered region (IDR) engages DNA and key repair proteins such as RAD51, BLM, and WRN. DNA binding through the BARD1 IDR facilitates H2A ubiquitination by the BRCA1-BARD1 complex and is essential for stimulating long-range DNA end resection during homologous recombination, underscoring its role in accurate DNA repair. Despite these insights, structural characterization of the IDR remains limited, leaving questions regarding its functional interplay with BRCA1 and other repair factors unresolved. Here, we report the backbone resonance assignments of a BARD1 IDR construct spanning residues 124-270, providing a foundation for future studies aimed at understanding how the disordered regions of BARD1 interact with various binding partners, and cooperates with itself and BRCA1 to regulate genome stability.
