Cytochrome <i>bd</i> Biosynthesis in <i>Bacillus subtilis</i> : Characterization of the <i>cydABCD</i> Operon

<jats:title>ABSTRACT</jats:title> <jats:p> Under aerobic conditions <jats:italic>Bacillus subtilis</jats:italic> utilizes a branched electron transport chain comprising various cytochromes and terminal oxidases. At present there is evidence for three types of terminal oxidases in <jats:italic>B. subtilis</jats:italic> : a <jats:italic>caa</jats:italic> <jats:sub>3</jats:sub> -, an <jats:italic>aa</jats:italic> <jats:sub>3</jats:sub> -, and a <jats:italic>bd</jats:italic> -type oxidase. We report here the cloning of the structural genes ( <jats:italic>cydA</jats:italic> and <jats:italic>cydB</jats:italic> ) encoding the cytochrome <jats:italic>bd</jats:italic> complex. Downstream of the structural genes, <jats:italic>cydC</jats:italic> and <jats:italic>cydD</jats:italic> are located. These genes encode proteins showing similarity to bacterial ATP-binding cassette (ABC)-type transporters. Analysis of isolated cell membranes showed that inactivation of <jats:italic>cydA</jats:italic> or deletion of <jats:italic>cydABCD</jats:italic> resulted in the loss of spectral features associated with cytochrome <jats:italic>bd</jats:italic> . Gene disruption experiments and complementation analysis showed that the <jats:italic>cydC</jats:italic> and <jats:italic>cydD</jats:italic> gene products are required for the expression of a functional cytochrome <jats:italic>bd</jats:italic> complex. Disruption of the <jats:italic>cyd</jats:italic> genes had no apparent effect on the growth of cells in broth or defined media. The expression of the <jats:italic>cydABCD</jats:italic> operon was investigated by Northern blot analysis and by transcriptional and translational <jats:italic>cyd-lacZ</jats:italic> fusions. Northern blot analysis confirmed that <jats:italic>cydABCD</jats:italic> is transcribed as a polycistronic message. The operon was found to be expressed maximally under conditions of low oxygen tension. </jats:p>