Role of Ca2+ 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 I50 of approximately 2.5 μM. The effect of Mg2+ and Ca2+ on EGTA inhibition of purified Complex I activity indicated that Ca2+ 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 Fe2+ or Zn2+. 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 Ca2+ binding site.

Research Highlights
► E. coli Complex I activity is strongly inhibited by divalent cation chelators.
► The extent of the inhibition is dependent on whether Complex I is membrane-bound or purified.
► The chelator's effect can be completely prevented by an equimolar amount of Ca2+ but not Mg2+.
► Complex I is suggested to contain tightly bound Ca2+ and the loss of this bound Ca2+ results in irreversible damage of the enzyme.