Effects of copper concentration in continuous culture on the aa 3-type cytochrome oxidase and respiratory chains of Paracoccus denitrificans

Abstract

The role(s) of copper in a bacterial cytochrome oxidase of the aa3-type was investigated by growth of Paracoccus denitrificans NCIB 8944, in batch and steady state continuous culture, in a medium from which the bulk of the copper had been extracted. In a medium containing approximately 0.02 μM copper, cellular copper content, cytochromes a+a3 and cytochrome a3 were reduced to 55%, 58% and 33% respectively of control values and there were also less marked decreases in cytochromes c+c1 (to 85%) and a CO-binding b-type cytochrome, possibly cytochrome o (to 71%). Copper deficiency elicited in reduced minus oxidized difference spectra a shift to shorter wavelengths and narrowing of the band width of the α-band of the oxidase, and loss of a (negative) band near 830 nm attributable to CuA (the copper functionally associated with haem a in the oxidase complex). The oxidase in copper-deficient cells reacted with oxygen to form the oxy “Compound A” at rates similar to that in control cells but CO recombination to ferrous haem a3 was slowed 4-fold in the copper deficient case. The results are interpreted as indicating loss of CuA and changes in the proportions of haems a and a3 with retention of catalytic activity. Titrations of respiration rates with antimycin suggested that copper deficiency did not result in diversion of electron flux through an antimycin A-insensitive, cytochrome o-terminated branch of the respiratory chain.