NAD(H) phosphates mediate tetramer assembly of human C-terminal binding protein (CtBP)
| Autor: | William E. Royer, Celia A. Schiffer, Jeffry C. Nichols |
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| Rok vydání: | 2021 |
| Předmět: |
Models
Molecular 0301 basic medicine Dehydrogenase Transcription coregulator Biochemistry DNA-binding protein MALS 03 medical and health sciences Tetramer tetrameric assembly TCEP tris(2-carboxyethyl) phosphine CtBP C-terminal binding protein medicine Humans structural biology Nucleotide CtBP crystallography Molecular Biology chemistry.chemical_classification 030102 biochemistry & molecular biology Chemistry NAD(H) MALS multiangle light scattering D2-HDH D-isomer-specific 2-hydroxyacid dehydrogenase Cell Biology NAD Adenosine DNA-Binding Proteins Alcohol Oxidoreductases transcription coregulator SEC size-exclusion column 030104 developmental biology Structural biology dehydrogenase Biophysics cancer target NAD+ kinase Protein Multimerization Co-Repressor Proteins NADP Research Article medicine.drug |
| Zdroj: | The Journal of Biological Chemistry |
| ISSN: | 0021-9258 |
| DOI: | 10.1016/j.jbc.2021.100351 |
| Popis: | C-terminal binding proteins (CtBPs) are cotranscriptional factors that play key roles in cell fate. We have previously shown that NAD(H) promotes the assembly of similar tetramers from either human CtBP1 and CtBP2 and that CtBP2 tetramer destabilizing mutants are defective for oncogenic activity. To assist structure-based design efforts for compounds that disrupt CtBP tetramerization, it is essential to understand how NAD(H) triggers tetramer assembly. Here, we investigate the moieties within NAD(H) that are responsible for triggering tetramer formation. Using multiangle light scattering (MALS), we show that ADP is able to promote tetramer formation of both CtBP1 and CtBP2, whereas AMP promotes tetramer assembly of CtBP1, but not CtBP2. Other NAD(H) moieties that lack the adenosine phosphate, including adenosine and those incorporating nicotinamide, all fail to promote tetramer assembly. Our crystal structures of CtBP1 with AMP reveal participation of the adenosine phosphate in the tetrameric interface, pinpointing its central role in NAD(H)-linked assembly. CtBP1 and CtBP2 have overlapping but unique roles, suggesting that a detailed understanding of their unique structural properties might have utility in the design of paralog-specific inhibitors. We investigated the different responses to AMP through a series of site-directed mutants at 13 positions. These mutations reveal a central role for a hinge segment, which we term the 120s hinge that connects the substrate with coenzyme-binding domains and influences nucleotide binding and tetramer assembly. Our results provide insight into suitable pockets to explore in structure-based drug design to interfere with cotranscriptional activity of CtBP in cancer. |
| Databáze: | OpenAIRE |
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