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All tied up: Tethered protein provides long-sought answer
Date:9/22/2009

of DNA. The combination of protein binding and the loop prevent the machinery that encodes proteins from copying the DNA, so in essence, lactose repressor "turns off" the nearby genes.

Lactose repressor draws its name from the genes that it blocks -- genes that encode enzymes used to transport and metabolize lactose in bacteria. If a bacterial cell happens to be where lactose is plentiful, lactose repressor binds to a derivative of lactose that prevents high-affinity binding of repressor to the DNA. The cell is then able to manufacture the enzymes needed to convert the lactose into food. If no lactose is present, the protein clamps onto the DNA and inactivates the process of copying the lactose genes so the cell doesn't waste energy making the enzyme.

In 2007, the University of Florence's Francesco Vanzi visited Matthews' lab to learn new techniques for purifying and assaying samples of lactose repressor. The protein has a limited shelf life, and Vanzi, who was preparing to do single-molecule studies on the protein, needed to find out how to make it on-site in his lab.

"While he was here, we talked about various ways to fix the two arms of the protein with cross-linkers," Matthews said. The idea was to bind the arms together with chemical manacles that would limit the movement around the hinge of the "V." Vanzi, Matthews and Rice postdoctoral fellow Hongli Zhan wound up using three sets of manacles, or tethers, including longer and shorter chemical tethers; they also used some reversible tethers to allow return to the protein's original state. The researchers chose two different binding sites, one that provided some degree of flexibility in opening the structure and one that kept the arms bound in the more-closed "V" position characteristic of the structure determined for the protein in crystals.

The team found that the more they restricted the flexibility of the arms, the less likely the protein was to create DNA loops by binding
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Contact: Jade Boyd
jadeboyd@rice.edu
713-348-6778
Rice University
Source:Eurekalert  

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