The Isolation of ASPP2-interacting Proteins with the Yeast-two Hybrid Assay

The Isolation of ASPP2-interacting Proteins with the Yeast-two Hybrid Assay

4.11 - 1251 ratings - Source

The protein ASPP2 is a major stimulator of p53, a tumor-suppressor protein that can activate and repress transcription. Probably the single most frequent genetic abnormality in human cancer is p53 mutations. The ability of p53 to induce apoptosis is a key defense against cancer. Regulators of p53 are partially to blame for the breakdown in the defense against cancer, which makes their identification and characterization essential. The p53 binding proteins, termed p53BP2 and Bbp are both fragments of ASPP2. It has been shown that the C-terminal fragment of ASPP2 interacts with several proteins that modulate its role in regulating apoptosis and p53 activity. Since ASPP2-interacting proteins were identified using the interacting proteins as bait to screen cDNA libraries, there is the possibility that other ASPP2-interacting proteins exist, which inhibits or stimulates the activity of ASPP2. In this thesis a human skeletal muscle library was screened with a yeast two-hybrid assay to identify additional proteins that interact with ASPP2 at its C-terminal and N-terminal regions. Several positive clones were isolated under medium/high stringency conditions, retested individually with the respective bait vector for alpha or beta galactosidase activation and sequenced for identification. Fragments of cDNA that encode several polypeptides were isolated, e.g., telethonin, actin, prohibitin and proteins of unknown function. The identification of these putative ASPP2-binding proteins will possibly increase the current knowledge of the ASPP2's p53-stimulatory function and reveal other pathways that may be involved in carcinogenesis.MATCHMAKER Gal4 Two-Hybrid system 3 aamp; Libraries User Manual 21. BD YeastmakerTM ... Apoptosis and syncytial fusion in human placental trophoblast and skeletal muscle. Int Rev Cytol ... iASPP preferentially binds p53 proline-rich region and modulates apoptotic function of codon 72-polymorphicp53. Nat. Geneticsanbsp;...

Title:The Isolation of ASPP2-interacting Proteins with the Yeast-two Hybrid Assay
Publisher:ProQuest - 2007


You Must CONTINUE and create a free account to access unlimited downloads & streaming