Role of Ca(2+) in structure and function of Complex I from Escherichia coli.
The dependence of E. coli Complex I activity on cation chelators such as EDTA, EGTA, NTA and o-phenanthroline was studied in bacterial membranes, purified solubilized enzyme and Complex I reconstituted into liposomes. Purified Complex I was strongly inhibited by EDTA with an I(50) of approximately 2.5μM. The effect of Mg(2+) and Ca(2+) on EGTA inhibition of purified Complex I activity indicated that Ca(2+) is tightly bound to the enzyme and essential for the activity. Low sensitivity to o-phenanthroline argues against the occupation of this cation binding site by Fe(2+) or Zn(2+). The sensitivity of Complex I to EDTA/EGTA strongly depends on the presence of monovalent cations in the medium, and on whether the complex is native, membrane-bound, or purified. The data is discussed in terms of a possible loss either of an additional 14th, subunit of E. coli Complex I, analogous to Nqo15 in the T. thermophilus enzyme, or another component of the native membrane that affects the affinity and/or accessibility of the Ca(2+) binding site.
Verkhovskaya
Marina
M
Helsinki Bioenergetics Group, Institute of Biotechnology, University of Helsinki, Finland. Marina.Verhovskaya@Helsinki.Fi
Knuuti
Juho
J
Wikström
Mårten
M
D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D002462:Cell Membrane / Q000201:enzymology; D004492:Edetic Acid / Q000494:pharmacology; D004533:Egtazic Acid / Q000494:pharmacology; D042967:Electron Transport Complex I / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology* / Q000254:growth & development; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D009243:NAD / Q000378:metabolism; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D012995:Solubility; D016963:Thermus thermophilus / Q000201:enzymology
D021122:Protein Subunits; D009243:NAD; D004533:Egtazic Acid; D004492:Edetic Acid; D042967:Electron Transport Complex I; D008274:Magnesium; D002118:Calcium