Binding Site Determinants for the LysR-Type Transcriptional Regulator PcaQ in the Legume Endosymbiont
Sinorhizobium melilotiJournal Articles
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ABSTRACT
LysR-type transcriptional regulators represent one of the largest groups of prokaryotic regulators described to date. In the gram-negative legume endosymbiont
Sinorhizobium meliloti
, enzymes involved in the protocatechuate branch of the β-ketoadipate pathway are encoded within the
pcaDCHGB
operon, which is subject to regulation by the LysR-type protein PcaQ. In this work, purified PcaQ was shown to bind strongly (equilibrium dissociation constant, 0.54 nM) to a region at positions −78 to −45 upstream of the
pcaD
transcriptional start site. Within this region, we defined a PcaQ binding site with dyad symmetry that is required for regulation of
pcaD
expression in vivo and for binding of PcaQ in vitro. We also demonstrated that PcaQ participates in negative autoregulation by monitoring expression of
pcaQ
via a transcriptional fusion to
lacZ
. Although
pcaQ
homologues are present in many α-proteobacteria, this work describes the first reported purification of this regulator, as well as characterization of its binding site, which is conserved in
Agrobacterium tumefaciens
,
Rhizobium leguminosarum
,
Rhizobium etli
, and
Mesorhizobium loti
.
