Terminal oxidases of Escherichia coli aerobic respiratory chain. I. Purification and properties of cytochrome b562-o complex from cells in the early exponential phase of aerobic growth.
Cytochrome b562-o complex, a terminal oxidase in the respiratory chain of aerobically grown Escherichia coli K12, was isolated in a highly purified form. The purified oxidase is composed of equimolar amounts of two polypeptides, with Mr = 33,000 and 55,000, determined by gel electrophoresis in the presence of sodium dodecyl sulfate. It contains 19.5 nmol of heme and 16.8 nmol of copper/mg of protein, but no detectable nonheme iron, phospholipid, ubiquinone, or menaquinone. In the difference spectrum at room temperature, the oxidase shows a single alpha absorption peak at 560 nm and at 77 K it shows two alpha absorption peaks at 555 and 562 nm. This oxidase combines with CO and the CO difference spectrum at room temperature has a peak at 416 nm and a trough at 430 nm in the Soret region. Its oxidation-reduction potential is estimated to be 125 mV (pH 7.4) and it is pH-dependent (-60 mV/pH) in medium of pH 6.0 to 7.4. It catalyzes electron transport to oxygen via ubiquinol and ascorbate in the presence of phenazine methosulfate or N,N,N',N'-tetramethyl-p-phenylenediamine dihydrochloride. This oxidase activity depends on phospholipids and is sensitive to respiratory inhibitors, such as 2-heptyl-4-hydroxyquinoline N-oxide, piericidin A, KCN and NaN3. The divalent cations Zn2+, Cd2+, and Co2+ inhibit the oxidase activity extensively. The oxidase activity of the cytochrome b562-o complex was inhibited by photoinactivation with rose bengal, suggesting that the inhibition by zinc ion results from modification of a histidine residue of cytochrome o.
D000332:Aerobiosis; D002248:Carbon Monoxide; D003300:Copper / Q000032:analysis; D003573:Cytochrome b Group / Q000302:isolation & purification* / Q000378:metabolism; D003580:Cytochromes / Q000302:isolation & purification* / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins; D006418:Heme / Q000032:analysis; D007501:Iron / Q000032:analysis; D007700:Kinetics; D008970:Molecular Weight; D009097:Multienzyme Complexes / Q000302:isolation & purification* / Q000378:metabolism; D010101:Oxygen Consumption; D010743:Phospholipids / Q000494:pharmacology; D011808:Quinone Reductases / Q000378:metabolism; D013053:Spectrophotometry
D003573:Cytochrome b Group; D003580:Cytochromes; D029968:Escherichia coli Proteins; D009097:Multienzyme Complexes; D010743:Phospholipids; D006418:Heme; D003300:Copper; D002248:Carbon Monoxide; C017168:cytochrome bo, E coli; C017850:cytochrome b562, E coli; D007501:Iron; D011808:Quinone Reductases; C030914:ubiquinol-1 oxidase