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Papers In Press, published online ahead of print November 20, 2001
J. Biol. Chem, 10.1074/jbc.M110555200
Submitted on November 2, 2001
Revised on November 20, 2001
Accepted on November 19, 2001

Constitutive mutations of the OccR regulatory protein affect DNA bending in response to metabolites released from plant tumors

Reiko Akakura and Stephen C. Winans

Microbiology, Cornell University, Ithaca, NY 14853

Corresponding Author: scw2{at}cornell.edu

OccR is a LysR-type transcriptional regulator of Agrobacterium tumefaciens that positively regulates the octopine catabolism operon of the Ti plasmid and is also an autorepressor. Positive control of the occ genes occurs in response to octopine, a nutrient released from crown gall tumors. OccR binds to a site upstream of the occQ promoter in the presence and absence of octopine. Octopine causes prebound OccR to undergo a conformational change at the DNA binding site that causes changes in footprint length and DNA bending. In order to determine the roles of these conformational changes in transcriptional activation, we isolated 22 OccR mutants that were able to activate the occQ promoter in the absence of octopine. Thirteen of these mutants contained single amino acid substitutions while nine contained two base pair changes resulting in two amino acid substitutions, which in most cases acted synergistically. These mutations spanned the entire length of the protein. Most of these mutant proteins, in the absence of octopine, displayed DNA binding and bending properties characteristic of transcriptionally active OccR-octopine complexes.


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