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| Content Provider | World Health Organization (WHO)-Global Index Medicus |
|---|---|
| Author | Shapleigh, J. Schmidt, E. Ferguson-miller, S. García-horsman, A. Fetter, J. R. Hosler, J. Babcock, G. T. Thomas, J. W. Qian, J. Gennis, R. B. |
| Description | Author Affiliation: Fetter JR ( Department of Biochemistry, Michigan State University, East Lansing 48824.); |
| Abstract | As the final electron acceptor in the respiratory chain of eukaryotic and many prokaryotic organisms, cytochrome c oxidase (EC 1.9.3.1) catalyzes the reduction of oxygen to water and generates a proton gradient. To test for proton pathways through the oxidase, site-directed mutagenesis was applied to subunit I of the Rhodobacter sphaeroides enzyme. Mutants were characterized in three highly conserved regions of the peptide, comprising possible proton loading, unloading, and transfer sites: an interior loop between helices II and III (Asp132Asn/Ala), an exterior loop between helices IX and X (His411Ala, Asp412Asn, Thr413Asn, Tyr414Phe), and the predicted transmembrane helix VIII (Thr352Ala, Pro358Ala, Thr359Ala, Lys362Met). Most of the mutants had lower activity than wild type, but only mutants at residue 132 lost proton pumping while retaining electron transfer activity. Although electron transfer was substantially inhibited, no major structural alteration appears to have occurred in D132 mutants, since resonance Raman and visible absorbance spectra were normal. However, lower CO binding (70-85% of wild type) suggests some minor change to the binuclear center. In addition, the activity of the reconstituted Asp132 mutants was inhibited rather than stimulated by ionophores or uncoupler. The inhibition was not observed with the purified enzyme and a direct pH effect was ruled out, suggesting an altered response to the electrical or pH gradient. The results support an important role for the conserved II-III loop in the proton pumping process and are consistent with the possibility of involvement of residues in helix VIII and the IX-X loop. |
| ISSN | 00278424 |
| e-ISSN | 10916490 |
| Journal | Proceedings of the National Academy of Sciences of the United States of America |
| Issue Number | 5 |
| Volume Number | 92 |
| Language | English |
| Publisher | National Academy of Sciences |
| Publisher Date | 1995-04-01 |
| Publisher Place | United States |
| Access Restriction | Open |
| Subject Keyword | Electron Transport Complex IV Metabolism Proton-Translocating ATPases Amino Acid Sequence Bacterial Proteins Chemistry Binding Sites Carbonyl Cyanide M-Chlorophenyl Hydrazone Pharmacology Electron Transport Hydrogen-Ion Concentration Molecular Sequence Data Mutagenesis, Site-Directed Rhodobacter Sphaeroides Enzymology Spectrum Analysis, Raman Structure-Activity Relationship Valinomycin Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. Multidisciplinary |
| Content Type | Text |
| Resource Type | Article |
| Subject | Multidisciplinary |
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