CLONING AND CHARACTERIZATION OF BTR, A BORDETELLA-PERTUSSIS GENE ENCODING AN FNR-LIKE TRANSCRIPTIONAL REGULATOR

To determine whether hemolytic factors other than the bifunctional hem olysin-adenylate cyclase toxin (cyclolysin) are expressed by Bordetell a pertussis, a gene library was constructed from a virulent strain of B. pertussis, BP504, transformed into nonhemolytic Escherichia coli, a nd screened on blood agar plates. A strongly hemolytic colony which co ntained the plasmid pHLY1A was isolated. Nucleotide sequencing of pHLY 1A revealed an open reading frame that could encode a 27-kDa protein. No similarity was detected between the deduced amino acid sequence of this open reading frame and those of any known bacterial cytolysins. H owever, significant homology was detected with FNR of E. coli and seve ral other transcriptional regulators including HlyX from Actinobacillu s pleuropneumoniae, which can also confer a hemolytic phenotype on E. coli. An fnr mutant of E. coli, JRG1728, could be complemented by pHLY 1A. Thus, the B. pertussis transcriptional regulator-like gene and the protein which it encoded were named btr and BTR, respectively. A BTR- deficient B. pertussis strain, BJB1, was constructed. The btr::kan mut ation had no effect on the expression of hemolytic activity or on phas e variation. Northern (RNA) blotting revealed that btr expression was not regulated by the BvgAS two-component sensor-regulator. On the basi s of sequence similarity to FNR-like transcriptional regulators and th e ability to complement an anaerobically deficient E. coli strain (JRG 1728) in growing anaerobically, BTR may regulate B. pertussis gene exp ression in response to changes in oxygen levels or to changes in the r edox potential of the bacterial environment. Its role in virulence rem ains to be determined.