John J. Hill

Insight into the active-site structure and function of cytochrome oxidase by analysis of site-directed mutants of bacterial cytochrome aa3 and cytochrome bo

Cytochromeaa3 ofRhodobacter sphaeroides and cytochromebo ofE. coli are useful models of the more complex cytochromec oxidase of eukaryotes, as demonstrated by the genetic, spectroscopic, and functional studies reviewed here. A summary of site-directed mutants of conserved residues in these two enzymes is presented and discussed in terms of a current model of the structure of the metal centers and evidence for regions of the protein likely to be involved in proton transfer. The model of ligation of the hemea3 (oro)-CuB center, in which both hemes are bound to helix X of subunit I, has important implications for the pathways and control of electron transfer.

Recent studies of the cytochrome o terminal oxidase complex of Escherichia coli

The cytochrome o complex is the predominant terminal oxidase in the aerobic respiratory chain of Escherichia coli when the bacteria are grown under conditions of high aeration. The oxidase is a ubiquinol oxidase and reduces molecular oxygen to water. Electron transport through the enzyme is coupled to the generation of a protonmotive force. The purified cytochrome o complex contains four or five subunits, two protoheme IX (heme b) prosthetic groups, plus at least one Cu. The subunits are all encoded by the cyo operon. Sequence comparisons show that the cytochrome o complex is closely related to the aa3-type cytochrome c oxidase family. Gene fusions have been used to define the topology of each of the gene products. Subunits I, II, III and IV are proposed to have 15, 2, 5 and 3 transmembrane spans, respectively. The fifth gene product (cyoE) encodes a protein with 7 membrane spanning segments, and this may also be a subunit of this enzyme. Fourier transform infrared spectroscopy has been used to monitor CO bound in the active site where oxygen is reduced. These data provide definitive proof that the cytochrome o complex has a heme-copper binuclear center, similar to that present in the aa3-type cytochrome c oxidases. Site-directed mutagenesis is being utilized to define which amino acids are ligands to the heme iron and copper prosthetic groups.

The value of history and goals of care with code status; even in an emergency setting

This is a case report involving an elderlyJehovahs Witness woman with a history of chronic anaemia who suffers cardiac arrest and is resuscitated. This case illustrates, even in an emergency setting, (1) the value of obtaining relevant history, (2) addressing code status within the framework of achievable goals of care and (3) the need to revise both history and goals of care, as more information becomes available. We also propose aspects of this discussion that would benefit from future research.

Photoselective discrimination of matrix isolated respiratory enzymes from E. coli by Fourier transform infrared spectroscopy

Photodissociation of heme-CO complexes in bacterial membranes was measured by Fourier transform infrared spectroscopy. Wavelength of the photodissociating light and selected cryogenic temperatures were used to distinguish multiple forms of respiratory enzymes (cytochromes).

Instrumental Considerations In Infrared Biospectroscopy

The 1959 cm-1 KBr impurity absorption in beamsplitters provides a convenient measurement of instability that may be used in conjunction with interferogram fluctuations to assess the total instrumental instability. Analysis of interferogram fluctuations led to environmental controls which reduced the KBr impurity band contribution in a ratioed spectrum to less than 0.9%, and facilitated measurement of sub-milliabsorbance infrared bands.