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[Identification of a protein interacting with apoptin from human leucocyte cDNA library by using yeast two-hybrid
1Institute of Radiation Medicine, Academy of Military Medical Sciences, Beijing 100850, China. sunzx@nic.bmi.ac.cn
Abstract:
To screen the protein interacting with apoptin from human leucocyte cDNA library by using yeast two-hybrid system, four clones interacting with apoptin were identified. One of them was homologous with Nmi (N-Myc interaction protein). Cell co-immunoprecipitation showed that apoptin could bind to Nmi in mammalian cells. Apoptin mutants T1, T2 and T3 lacked the C-terminal 11 AA,33-46 AA and both,respectively. Apoptin mutants T2 and T3 failed to interact with Nmi, suggesting that its 33-46 AA was pivotal for the interaction. Apoptin mutant T1 still interacted with Nmi, suggesting that its C-terminal 11 AA was not essential for the interaction.
Insights
Researchers identified N-Myc interaction protein (Nmi) as a binding partner of apoptin using a yeast two-hybrid system. The study found that the 33-46 amino acid region of apoptin is crucial for this interaction.
Area of Science:
- Molecular Biology
- Cell Biology
- Protein-Protein Interactions
Background:
- Apoptin is a protein with tumor-killing properties.
- Understanding apoptin's interactions is key to its therapeutic potential.
Purpose of the Study:
- To identify proteins that interact with apoptin.
- To determine the specific region of apoptin involved in binding to its interacting partners.
Main Methods:
- Yeast two-hybrid screening of a human leucocyte cDNA library.
- Co-immunoprecipitation assays in mammalian cells.
- Analysis of apoptin mutants with specific amino acid deletions.
Main Results:
- Four proteins interacting with apoptin were identified, including N-Myc interaction protein (Nmi).
- Apoptin directly binds to Nmi in mammalian cells.
- The 33-46 amino acid region of apoptin is essential for Nmi interaction, while the C-terminal 11 amino acids are not.
Conclusions:
- Nmi is a novel binding partner of apoptin.
- The 33-46 amino acid domain of apoptin plays a critical role in mediating its interaction with Nmi.
- This interaction may be important for apoptin's biological functions.