Cholate-induced dimerization of detergent- or phospholipid-solubilized bovine cytochrome C oxidase.
Bovine heart cytochrome c oxidase (CcO), solubilized by either nonionic detergents or phospholipids, completely dimerizes upon the addition of bile salts, e.g., sodium cholate, sodium deoxycholate, or CHAPS. Bile salt induced dimerization occurs whether dodecyl maltoside, decyl maltoside, or Triton X-100 is the primary solubilizing detergent or the enzyme is dispersed in phosphatidylcholine, phosphatidylethanolamine, or mixtures thereof. In each case, complete CcO dimerization can be verified by sedimentation velocity and sedimentation equilibrium after correction for bound detergent and/or phospholipid. The relative concentration of the bile salt is critical for production of homogeneous, dimeric CcO. For example, enzyme solubilized by 2 mM detergent requires an equal molar concentration of sodium cholate. Similarly, enzyme dispersed in 20 mM phospholipid requires 50 mM sodium cholate, concentrations that are commonly used to reconstitute CcO into small unilamellar vesicles. Bile salts do more than just stabilize dimeric CcO and prevent detergent-induced dissociation into monomers. They are able to completely reverse detergent-induced monomerization and cause completely monomeric CcO to reassociate. Dimeric CcO so generated is no more stable than the original complex and easily dissociates into monomers if the bile salt is removed. The dimerization process is dependent upon a full complement of subunits; e.g., if subunits VIa and VIb are removed, the resulting monomeric CcO will not reassociate upon the addition of sodium cholate. These results support four important consequences: (1) dissociation of dimeric CcO into monomers is reversible; (2) stable dimers can be produced under solution conditions; (3) dimers can be stabilized even at relatively high pH and low enzyme concentration; and (4) subunits VIa and VIb are required for dimerization.
Musatov
Andrej
A
Department of Biochemistry, The University of Texas Health Science Center, 7703 Floyd Curl Drive, San Antonio, Texas 78229-3900, USA.
Robinson
Neal C
NC
D000818:Animals; D001647:Bile Acids and Salts / Q000494:pharmacology; D002417:Cattle; D020355:Cholates / Q000494:pharmacology*; D002793:Cholic Acids / Q000494:pharmacology; D002852:Chromatography, Ion Exchange; D003840:Deoxycholic Acid / Q000494:pharmacology; D003902:Detergents / Q000494:pharmacology*; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D003576:Electron Transport Complex IV / Q000737:chemistry / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008823:Micelles; D009206:Myocardium / Q000737:chemistry; D010743:Phospholipids / Q000494:pharmacology*; D011485:Protein Binding; D017434:Protein Structure, Tertiary
D001647:Bile Acids and Salts; D020355:Cholates; D002793:Cholic Acids; D003902:Detergents; D008823:Micelles; D010743:Phospholipids; D003840:Deoxycholic Acid; D003576:Electron Transport Complex IV; C028213:3-((3-cholamidopropyl)dimethylammonium)-1-propanesulfonate