The respiratory complex I (NDH I) from Klebsiella pneumoniae, a sodium pump.
The electrogenic NADH:Q oxidoreductase from the enterobacterium Klebsiella pneumoniae transports Na(+) ions. The complex was purified with an increase of the specific Na(+) transport activity from 0.2 micromol min(-1) mg(-1) in native membrane vesicles to 4.7 micromol min(-1) mg(-1) in reconstituted enzyme specimens. The subunit pattern resembled that of complex I from Escherichia coli, and two prominent polypeptides were identified as the NuoF and NuoG subunits of complex I. During purification the typical cofactors of complex I were enriched to yield approximately 17 nmol mg(-1) iron, 24 nmol mg(-1) acid-labile sulfide, and 0.79 nmol mg(-1) FMN in the purified sample. The enzyme contained approximately 1.2 nmol mg(-1) Q6 and 1.5 nmol mg(-1) Q8. The reduction of ubiquinone by NADH was Na(+)-dependent, which indicates coupling of the chemical and the vectorial reaction of the pump. The Na(+) activation profile corresponded to the Hill equation with a Hill coefficient K(H)(Na(+)) = 1.96 and with a half-maximal saturation at 0.33 mm Na(+). The reconstituted complex I from Klebsiella pneumoniae catalyzed deamino-NADH oxidation, Q1 reduction, and Na(+) translocation with specific activities of 2.6 units mg(-1), 2.4 units mg(-1), and 4.7 units mg(-1), respectively, which indicate a Na(+)/electron stoichiometry of one.
Mikrobiologisches Institut der Eidgenössischen Technischen Hochschule, ETH-Zentrum, Schmelzbergstrasse 7, CH-8092 Zürich, Switzerland.
Anja C
AC
Gemperli
Peter
P
Dimroth
Julia
J
Steuber
D017136:Ion Transport; D007700:Kinetics; D007711:Klebsiella pneumoniae / Q000201:enzymology*; D010084:Oxidation-Reduction; D021122:Protein Subunits; D011808:Quinone Reductases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D012964:Sodium / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000302:isolation & purification*
D021122:Protein Subunits; D012964:Sodium; C041831:NADH dehydrogenase (quinone); D011808:Quinone Reductases; D000254:Sodium-Potassium-Exchanging ATPase