Specific modification of a Na+ binding site in NADH:quinone oxidoreductase from Klebsiella pneumoniae with dicyclohexylcarbodiimide.
The respiratory NADH:quinone oxidoreductase (complex I) (NDH-1) is a multisubunit enzyme that translocates protons (or in some cases Na+) across energy-conserving membranes from bacteria or mitochondria. We studied the reaction of the Na+-translocating complex I from the enterobacterium Klebsiella pneumoniae with N,N'-dicyclohexylcarbodiimide (DCCD), with the aim of identifying a subunit critical for Na+ binding. At low Na+ concentrations (0.6 mM), DCCD inhibited both quinone reduction and Na+ transport by NDH-1 concurrent with the covalent modification of a 30-kDa polypeptide. In the presence of 50 mM Na+, NDH-1 was protected from inhibition by DCCD, and the modification of the 30-kDa polypeptide with [14C]DCCD was prevented, indicating that Na+ and DCCD competed for the binding to a critical carboxyl group in NDH-1. The 30-kDa polypeptide was assigned to NuoH, the homologue of the ND1 subunit from mitochondrial complex I. It is proposed that Na+ binds to the NuoH subunit during NADH-driven Na+ transport by NDH-1.
Vgenopoulou
Irini
I
Biochemisches Institut, Universität Zürich, Winterthurerstr. 190, CH-8057 Zürich, Switzerland.
Gemperli
Anja C
AC
Steuber
Julia
J
D001665:Binding Sites / Q000187:drug effects; D004024:Dicyclohexylcarbodiimide / Q000494:pharmacology*; D007711:Klebsiella pneumoniae / Q000201:enzymology* / Q000378:metabolism; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism*
D004024:Dicyclohexylcarbodiimide; D012964:Sodium; D016660:NAD(P)H Dehydrogenase (Quinone)