Specificity and Binding Affinity of Phospholipids to the High-Affinity Cardiolipin Sites of Beef Heart Cytochrome c Oxidase

Neal C. Robinson

Research output: Contribution to journalArticle

83 Scopus citations

Abstract

Beef heart lipid-depleted cytochrome c oxidase, containing only 50% of the two to three essential high-affinity cardiolipin molecules per heme aa3 complex, was used to study the phospholipid requirements of this enzyme. The lipid-depleted complex had two-thirds of the electron transport activity as enzyme containing a full complement of essential cardiolipin (diphosphatidylglycerol or DPG) when it was assayed in Tween 80. However, incubation of the lipid-depleted enzyme with DPG in the presence of 1% Triton X-100 followed by a 140- fold dilution of the reconstituted complex into Tween 80 restored 100% of the initial activity. Similar incubations of the lipid-depleted enzyme with phosphatidylethanolamine, phosphatidylglycerol, phosphatidylserine, or phosphatidic acid did not stimulate the enzymatic activity in Tween 80 more than 5-10%. In the presence of 1% Triton X-100, bovine DPG (90% Cl8:2) reassociated with the vacant high-affinity sites with an apparent dissociation constant of 5 µ based upon the regeneration of electron transport activity in Tween 80. Bacterial DPG (0% C18:2) measured in a similar manner had a dissociation constant of 2 µM therefore, the binding of exogenous DPG to the high-affinity sites on the enzyme does not have an absolute specificity for the linoleic acid tails present in bovine DPG. Reisolation of the complex by discontinuous glycerol gradient centrifugation after incubation with exogenous DPG in 1% Triton X-100 indicated that DPG reassociated only with the high-affinity cardiolipin sites; phosphatidylcholine did not bind to the complex under these conditions. On the basis of these results, we conclude that the unique structure of DPG, not the negatively charged head group or the composition of the hydrocarbon tails, is the structural feature necessary for the binding of exogenous phospholipids to the essential high-affinity sites of cytochrome c oxidase and the maintenance of full electron transport activity in Tween 80.

Original languageEnglish (US)
Pages (from-to)184-188
Number of pages5
JournalBiochemistry
Volume21
Issue number1
DOIs
StatePublished - Jan 1982

ASJC Scopus subject areas

  • Biochemistry

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