Octopine is released from crown gall tumors as a nutrient source and a signal molecule for the plant pathogen Agrobacterium tumefaciens. Some or all octopine-inducible genes are regulated by a protein called OccR. Primer extension analysis showed that OccR protein represses the occR gene and both represses and activates the occQ operon, which is divergently transcribed from occR. These promoters initiate transcription 46 bp apart. This regulatory system was reconstituted in vitro using purified OccR protein and Escherichia coli RNA polymerase. OccR binds with high affinity to a single site overlapping these promoters. Octopine shortens the DNAase I footprint of OccR and increases the gel mobility of OccR-DNA complexes by relaxing an OccR-incited DNA bend.