System Of Two Hybrid Proteins example essay topic

Raj i cells Cell line that grows in suspension derived from patient with Burkitt&'s lymphoma lymphoblast Often referred to as a blast cell. Unlike other usages of the suffix -blast a lymphoblast is a further differentiation of a lymphocyte, T- or B-, occasioned by an antigenic stimulus. The lymphoblast usually develops by enlargement of a lymphocyte, active re-entry to the S phase of the cell cycle, mito genesis and production of much m-RNA and ribosomes. Burkitt's lymphoma Malignant tumour of lymphoblasts derived from B-lymphocytes.

Most commonly affects children in tropical Africa: both Epstein-Barr virus and immunosuppression due to malarial infection are involved. yeast-2-hybrid interactions All two-hybrid screening systems rely on the fact that transcriptional activation and DNA-binding domains of transcription factors are modular in nature 6, 7. In these systems, the coding sequence for the DNA-binding domain of a transcription factor such as Gal 4 or LexA is fused to the cDNA of a protein of interest, termed the "bait". The fusion protein thus encoded tethers the bait to the promoter region of a reporter gene. A second fusion of a cDNA library with the coding sequence of a transcriptional activation domain is termed the "prey".

Functional reconstitution of transcription factor activity occurs upon association of the bait and prey protein domains. This interaction is then detected by expression of reporter genes that are dependent upon the bait's DNA-binding domain. Thus, the two-hybrid system is ideal for screening libraries for novel protein-protein interactions, and is also a powerful tool to isolate factors that disrupt or promote protein interactions. Its advantage is that an interaction of the bait with an unknown prey leads to direct identification of the cDNA that encodes the interacting protein, and its use in high-throughput screening applications allows the rapid and efficient screening of large numbers of potential interacting cDNAs or peptides. We have generated a novel genetic system to study these interactions by taking advantage of the properties of the GAL 4 protein of the yeast Saccharomyces cerevisiae. This protein is a transcriptional activator required for the expression of genes encoding enzymes of galactose utilization 1.

It consists of two separable and functionally essential domains: an N-terminal domain which binds to specific DNA sequences (UASG); and a C-terminal domain containing acidic regions, which is necessary to activate transcription 2, 3. We have generated a system of two hybrid proteins containing parts of GAL 4: the GAL 4 DNA-binding domain fused to a protein 'X' and a GAL 4 activating region fused to a protein 'Y'. If X and Y can form a protein-protein complex and reconstitute proximity of the GAL 4 domains, tran-scription of a gene regulated by UASG occurs. We have tested this system using two yeast proteins that are known to interact SNF 1 and SNF 4. High transcriptional activity is obtained only when both hybrids are present in a cell. This system may be applicable as a general method to identify proteins that interact with a known protein by the use of a simple galactose selection..