Iron-sulfur cluster N7 of the NADH:ubiquinone oxidoreductase (complex I) is essential for stability but not involved in electron transfer.
The NADH:ubiquinone oxidoreductase (complex I) from Escherichia coli is composed of 13 subunits called NuoA through NuoN. It catalyzes the electron transfer from NADH to ubiquinone by a chain of redox groups consisting of one FMN and seven iron-sulfur clusters. The function of the additional, nonconserved cluster N7 located on NuoG is not known. It has been speculated that it is not involved in electron transfer, due to its distance of more than 20 A from the electron transfer chain. Dithionite-reduced minus NADH-reduced EPR difference spectra of complex I and of a soluble fragment containing NuoG revealed for the first time the EPR spectrum of N7 in the complex. Individual mutation of the cysteines ligating this cluster to alanine led to a decreased amount of complex I in the membrane without affecting the electron transfer activity. Sucrose gradient centrifugation revealed that the complex from the C230A and C233A mutants decayed in detergent solution while the C237A and C265A mutant complex was stable. Cluster N7 was detectable in the latter mutants but with shifted g-values, indicating a different ligation of N7. Thus, N7 is essential for the stability of the complex but is not involved in electron transfer.
Pohl
Thomas
T
Institut für Organische Chemie und Biochemie, Albert-Ludwigs-Universität Freiburg, Albertstrasse 21, Chemiehochhaus, D-79104 Freiburg i. Br., Germany.
Bauer
Theresa
T
Dörner
Katerina
K
Stolpe
Stefan
S
Sell
Philipp
P
Zocher
Georg
G
Friedrich
Thorsten
T
D004578:Electron Spin Resonance Spectroscopy; D004579:Electron Transport; D042967:Electron Transport Complex I / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D007506:Iron-Sulfur Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D008958:Models, Molecular; D015394:Molecular Structure; D016297:Mutagenesis, Site-Directed; D009243:NAD; D010084:Oxidation-Reduction; D021122:Protein Subunits / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D013347:Subcellular Fractions / Q000201:enzymology
D029968:Escherichia coli Proteins; D007506:Iron-Sulfur Proteins; D021122:Protein Subunits; D009243:NAD; D042967:Electron Transport Complex I