Na+ translocation by the NADH:ubiquinone oxidoreductase (complex I) from Klebsiella pneumoniae.
Complex I is the site for electrons entering the respiratory chain and therefore of prime importance for the conservation of cell energy. It is generally accepted that the complex I-catalysed oxidation of NADH by ubiquinone is coupled specifically to proton translocation across the membrane. In variance to this view, we show here that complex I of Klebsiella pneumoniae operates as a primary Na+ pump. Membranes from Klebsiella pneumoniae catalysed Na+-stimulated electron transfer from NADH or deaminoNADH to ubiquinone-1 (0.1-0.2 micromol min-1 mg-1). Upon NADH or deaminoNADH oxidation, Na+ ions were transported into the lumen of inverted membrane vesicles. Rate and extent of Na+ transport were significantly enhanced by the uncoupler carbonylcyanide-m-chlorophenylhydrazone (CCCP) to values of approximately 0.2 micromol min-1 mg-1 protein. This characterizes the responsible enzyme as a primary Na+ pump. The uptake of sodium ions was severely inhibited by the complex I-specific inhibitor rotenone with deaminoNADH or NADH as substrate. N-terminal amino acid sequence analyses of the partially purified Na+-stimulated NADH:ubiquinone oxidoreductase from K. pneumoniae revealed that two polypeptides were highly similar to the NuoF and NuoG subunits from the H+-translocating NADH:ubiquinone oxidoreductases from enterobacteria.
Krebs
W
W
Mikrobiologisches Institut der Eidgenössischen Technischen Hochschule, ETH-Zentrum, Schmelzbergstr. 7, CH-8092 Zürich, Switzerland.
Steuber
J
J
Gemperli
A C
AC
Dimroth
P
P
D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism; D001692:Biological Transport; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D003902:Detergents; D004123:Dimethylamines; D004579:Electron Transport; D042967:Electron Transport Complex I; D007700:Kinetics; D007711:Klebsiella pneumoniae / Q000201:enzymology*; D009243:NAD / Q000378:metabolism; D009247:NADH, NADPH Oxidoreductases / Q000737:chemistry*; D010446:Peptide Fragments / Q000737:chemistry; D011189:Potassium Chloride / Q000494:pharmacology; D011522:Protons; D017421:Sequence Analysis; D012964:Sodium / Q000378:metabolism*; D012965:Sodium Chloride / Q000494:pharmacology; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism
D001425:Bacterial Outer Membrane Proteins; D003902:Detergents; D004123:Dimethylamines; D010446:Peptide Fragments; D011522:Protons; D009243:NAD; D012965:Sodium Chloride; C014518:dodecyldimethylamine oxide; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; D011189:Potassium Chloride; D012964:Sodium; D009247:NADH, NADPH Oxidoreductases; D042967:Electron Transport Complex I; D000254:Sodium-Potassium-Exchanging ATPase