The Escherichia coli multi-promoter region of the gapA gene ensures a high level

Question

The Escherichia coli multi-promoter region of the gapA gene ensures a high level of GAPDH (glyceraldehyde-3-phosphate dehydrogenase) production under various growth conditions. In the exponential phase of growth, gapA mRNAs are mainly initiated at the highly efficient gapA P1 promoter. In the present study, we used site-directed mutagenesis and chemical probing of the RPo (the open transcription complex) formed by EO7 holoenzyme associated with 70 RNAP (RNA polymerase) at promoter gapA P1 to show that this promoter is an extended -10 promoter that needs a -35 sequence for activity. The -35 sequence compensates for the presence of a suboptimal -10 hexamer. A tract of thymine residues in the spacer region, which is responsible for a DNA distortion, is also required for efficient activity. We present the first chemical probing of an RPo formed at a promoter needing both a -10 extension and a -35 sequence. It reveals a complex array of RNAP-DNA interactions. In agreement with the fact that an "A" (position -11) in the non-template strand is flipped out into a protein pocket in a previously studied RPo, we show the corresponding A residue in gapA P1 promoter is protected in RPo and is essential for activity. However, in contrast with some of the previous findings on RPo's formed at other promoters, the -12 A:T pair is opened. Strong contacts with RNAP occur both with the-35 sequence and the TG extension, so that the two domains, 2 and 4 may simultaneously contact the promoter DNA. RNAP-DNA interactions were also detected immediately downstream of the -35 hexamer and in a more distal upstream segment, reflecting a wrapping of RNAP by the core and upstream promoter DNA. Altogether, the data reveal that promoter gapA P1 is a very efficient promoter sharing common properties with both extended -10 and non extended -10 promoters 2 Based on the terminology and concepts we learned concerning bacterial promoters, dissect this abstract according to the statements I underlined and numbered (it is only necessary to interpret the underlined statements) I. Statement #1 refers to the ability of this gene promoter to produce high amounts of a. gene DNA 2. Statement #2 indicates that the gapA promoter is a promoter b. gene product c. gene mRNA d. enzyme catalysis a. unusual b. medium C. normal d. strong 3, Statement #3 acknowledges that and RNA polymerase normally occur in the cell as proteins. a. separate b. attached c. incomplete d. non-functional bp upstream from the 1 bp 4, Statement #4 indicates that an adenine nucleotide in the promoter is located transcription start site? a. not enough information is provided b. 21 bp c. 10 bp d. 11 bp 5. Statement #5 reveals a. two different species of sigma factors, 05, and 4, are required for RNA pol binding at this promoter b, these are the two "parts" of the overall sigma factor 70 protein structure involved in binding to the DNA c. two sigma factor species combine with to bind the dsDNA strand at this part of the promoter d, two separate sigma factors, 05, and 4, compete for binding sites at this promoter 70

Explanation / Answer

1.gene product

2. Strong

3. Seperate

4. 10 BP

5. Two different sigma factors are required for RNA polymerase binding at this promoter

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