Proteins in a family of transcription factors (DNA binding molecules that activa

Question

Proteins in a family of transcription factors (DNA binding molecules that activate transcription) have 2 distinct domains: activation domain (AD) and DNA binding domain (DBD). These are not steroid hormone receptors since they do not require hormone binding to do their job. While studying a transcription factor (XP) you discover that if you break these two domains apart and attach them to two separate molecules (Y and Z) the transcription factor would still function as long as Y and Z bind to each other.

Which of the following would probably be true about this system? Explain each briefly. (8)

A.The AD and DBD cannot truly be considered

Explanation / Answer

Hi,

Transcriptional activator proteins recruit the RNA polymerase II machinery and chromatin-modifying activities to promoters. Biochemical experiments indicate that activator proteins can associate with a large number of proteins, and many such proteins have been proposed to be direct targets of activators. However, there is great uncertainty about which biochemical interactions are physiologically relevant. Here, we develop a formaldehyde-based cross-linking procedure to identify protein-protein interactions that occur under physiological conditions. We show that the VP16 activation domain directly interacts with TATA-binding protein (TBP), TFIIB, and the SAGA histone acetylase complex in vivo.

Transcriptional activator proteins regulate the expression of eukaryotic genes in response to developmental and environmental cues. Such activator proteins contain a DNA-binding domain that recognizes specific promoter DNA sequences and a physically separate transcriptional activation region that stimulates mRNA initiation by RNA polymerase II . Activation domains are functionally autonomous; they function when fused to heterologous DNA-binding domains tethered at different positions in the promoter region. The best characterized activation domains are defined by short acidic regions that show little primary sequence homology . Mutational analysis indicates that acidic and hydrophobic residues within these domains contribute to functional activity, although individual residues make only a minor contribution . Acidic activation domains do not have a defined tertiary structure but become structured only upon specific interaction with another protein . Taken together, these observations indicate that acidic activation domains are surfaces used to mediate protein-protein interactions. It is presumed that other types of activation domains, such as those rich in glutamine or proline residues, function in a similar manner.

It is thought that the function of the zinc binding domain is for sequence specific DNA recognition. DNA primases make RNA primers which are then used for DNA synthesis. The placement of the RNA primers is not random, suggesting that they are placed on specific DNA sequences. Indeed, other DNA primases have been shown to recognize triplet sequences; the specific sequence recognized by B. stearothermophilus has not yet been identified. It has been shown that if the cystine residues that coordinate the zinc ion are mutated, the DNA primase stops functioning. This indicates that the zinc-binding domain does play a role in sequence recognition. In addition, the hydrophobic surface of the ? sheet, as well as the basic residues which are clustered primarily on one edge of the sheet, serve to attract single stranded DNA, further facilitating DNA binding.

Based on previous studies of DNA binding by DNA Primases, it is thought that DNA binds to the zinc-binding domain across the surface of the ? sheet, with the three nucleotides binding across three strands of the ? sheet. The positively charged residues in the sheet would be able to form contacts with the phosphates and the aromatic residues would form stacking interactions with the bases. This is the model of DNA binding by the ssDNA-binding domain of replication protein A (RPA). It is logical to assume that B. stearothermophilus

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