Detecting cooperative protein-DNA interactions and DNA loop formation by footprinting.
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Bacteriophage lambda↗
Biomedical subjects
A Hochschild
Publications and source records attributed to A Hochschild.
Interaction at a distance between lambda repressors disrupts gene activation.
The lambda repressor is an activator as well as a repressor of transcription. The activation function is blocked by interaction with another lambda repressor molecule bound upstream on the same DNA molecule. This example of negative control at a distance involves formation of a DNA loop.
Bacteriophage lambda↗
How lambda repressor and lambda Cro distinguish between OR1 and OR3.
Although lambda repressor and lambda Cro bind to the same six operators on the phage chromosome, the fine specificities of the two proteins differ: repressor binds more tightly to OR1 than to OR3, and vice versa for Cro. In this paper, we change base pairs in the operators and amino acids in the proteins to analyze the basis for these preferences. We find that these preferences are determined by residues 5 and 6 of the recognition helices of the two proteins and by the amino-terminal arm, in the case of repressor. We also find that the most important base pairs in the operator which enable repressor and Cro to discriminate between OR1 and OR3 are position 3 (for Cro) and positions 5 and 8 (for repressor). These and previous results show how repressor and Cro recognize and distinguish between two related operator sequences.
Amino Acid Sequence↗
Homologous interactions of lambda repressor and lambda Cro with the lambda operator.
Lambda repressor and lambda Cro bind to the same six sites on the phage chromosome but with different relative affinities. Nucleotides at certain positions in the operator are conserved in all sites, as are amino acids at certain positions in the recognition alpha-helices of repressor and Cro. Here we focus on one of the conserved amino acids, a serine found at position 2 of each recognition helix. We show that, contrary to a previous model, both serines contact the same conserved position in the operator, position 4. We suggest a simplified view of how repressor and Cro recognized similar operator sites but distinguish differently among them.
Amino Acid Sequence↗
Cooperative binding of lambda repressors to sites separated by integral turns of the DNA helix.
Lambda repressors bind cooperatively to adjacent pairs of operator sites. Here we show that repressors bind cooperatively to pairs of operator sites whose centers have been separated by five or six turns of the helix. No cooperativity is observed when the centers of these sites are on opposite sides of the DNA helix. Cooperativity depends upon the same part of the protein (the carboxyl domain) that mediates cooperativity when the sites are adjacent. As the repressors bind, the DNA between the sites becomes alternately sensitive and resistant to DNAase I cleavage at half turn intervals. We suggest that when repressors bind cooperatively to separated sites, the DNA forms a loop, thus allowing the two repressors to touch.
Base Sequence↗
Successful women in the sciences: an analysis of determinants. Making it: marginality and obstacles to minority consciousness.
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Female↗
DNA loops induced by cooperative binding of lambda repressor.
It has been shown by Hochschild and Ptashne that lambda repressors bind cooperatively to operator sites separated by five or six turns of the helix. Cooperative binding is not observed if the sites are separated by a nonintegral number of turns, unless a four-nucleotide gap is introduced into one of the strands between the two sites. These and other facts suggested that repressors at the separated sites touch each other, the DNA bending smoothly so as to accommodate the protein-protein interaction. Here we use electron microscopy to visualize the predicted protein-DNA complexes.
Binding Sites↗