Question

1. In DETAIL describe the assembly of the general transcription factors at a class II gene. Provide the important components of each complex, list the assembly in the correct order, and describe the function of each complex through generation of the open complex (10 pts).

2. Define the following terms, for proteins provide their function. Remember, providing an example is not a definition (1pt each).

a. Consensus sequence

b. RNaseH

c. Ribonucleoprotein complex

d. Transcriptional capacity

Answer 1

1a) The assembly of general transcription factors at Class II gene follows the following steps.

TATA binding protein (TBP), a subunit of TFIID (the largest GTF) binds to the promoter (TATA box), creating a sharp bend in the promoter DNA. Then the TBP-TFIIA interactions recruit TFIIA to the promoter.
TBP-TFIIB interactions recruit TFIIB to the promoter. RNA polymerase II and TFIIF assemble to form the Polymerase II complex. TFIIB helps the Pol II complex bind correctly.
TFIIE and TFIIH then bind to the complex and form the transcription preinitiation complex. TFIIA/B/E/H leave once RNA elongation begins. TFIID will stay until elongation is finished.
Subunits within TFIIH that have ATPase and helicase activity create negative superhelical tension in the DNA. This negative superhelical tension causes approximately one turn of DNA to unwind and form the transcription bubble.
The template strand of the transcription bubble engages with the RNA polymerase II active site, then RNA synthesis starts.

2a) A consensus sequence is an ideal promoter sequence in DNA - in E. coli, for example, two are found, a -35 sequence and a -10 sequence. The ideal promoter sequence - the consensus sequence - is never actually found in DNA, and a promoter's strength can be judged by it's similarity to the consensus sequence. The closer a promoter is to the ideal sequence, the stronger it will be and therefore the more mRNAwill be produced, which will lead to a greater yield of proteins. The -35 consensus sequence is TTGACA, and the -10 consensus sequence is TATAAT.

2b) Ribonuclease H (RnaseH) is a family of non-sequence-specific endonucleaseenzymes that catalyze the cleavage of RNA in an RNA/DNA substrate via a hydrolytic mechanism. Members of the RNase H family can be found in nearly all organisms, from bacteria to archaea to eukaryotes.

2c) Ribonucleoprotein (RNP) is a nucleoprotein that contains RNA, i.e. it is an association that combines a ribonucleic acid and an RNA-binding proteintogether. Such a combination can also be referred to as a protein-RNA complex. These complexes play an integral part in a number of important biological functions that include DNA replication, regulating gene expression and regulating the metabolism of RNA.

2d)Transcriptional activity refers to the binding and processive activity of RNA polymerase. RNA expression usually refers to the amount of mature transcripts but is often used interchangeably with transcriptional activity.