Cytochromes from Mycobacterium phlei: II. Ascorbate reduction of an isolated cytochrome ( a + a3) ( o) complex

Cytochrome a + a 3 and cytochrome o of a cytochrome ( a + a 3) ( o) complex isolated from Mycobacterium phlei could be differentiated on the basis of their response to inorganic phosphate and to pH. At low concentrations of phosphate over a pH range from 6.0 to 7.5, the rate of reduction of cytochromes a + a 3 was stimulated and the peaks for cytochrome a 3-CO in the carbon monoxide difference spectrum were more prominent than those of cytochrome o-CO. The reverse was true at pH 8.0 in that low concentrations of phosphate did not stimulate cytochromes a + a 3 reduction, but enhanced the appearance of cytochrome o-CO in the carbon monoxide difference spectrum. High concentrations of phosphate inhibited cytochromes a + a 3 reduction from pH 6.0 to 8.0. Due to the large size of the isolated complex, the effect of depolymerizing agents and detergents was tested. Stimulation of the rate of cytochromes a + a 3 reduction occurred in the presence of urea or guanidine-HCl. The CO difference spectrum obtained in the presence of guanidine-HCl showed the presence of cytochrome a 3-CO but not cytochrome o-CO while with increasing concentrations of urea, cytochrome o-CO was reduced in addition to cytochrome a 3-CO. Detergents such as sodium dodecyl sulfate or deoxycholate were found to inhibit the reduction of cytochromes a + a 3.