Specific cardiolipin binding interferes with labeling of sulfhydryl residues in the adenosine diphosphate/adenosine triphosphate carrier protein from beef heart mitochondria

Biochemistry. 1996 Dec 10;35(49):15784-90. doi: 10.1021/bi9610055.

Abstract

The interaction of cardiolipin with the isolated ADP/ATP carrier protein from beef heart mitochondria has been studied by means of the unmasking of a single cysteinyl residue, Cys56, which accompanies the conformational transition of the protein [Leblanc, P., & Clauser, H, (1972) FEBS Lett. 23, 107-113]. The unmasking was monitored by using the static fluorescence of the sulfhydryl reagent N-(1-pyrenyl)maleimide (PYM). The rate of PYM binding that was observed after initiation of the conformational transition by ADP was drastically reduced in the presence of cardiolipin (CL). Phospholipids other than CL were much less effective. It can be shown that the conformational transition and the binding reaction are both affected by CL, although to varying extents. An enhancement of the rate of the ADP-dependent PYM binding was observed upon digestion of the protein bound phospholipid by phospholipase A2. The phospholipase treatment also led to an increased ADP-independent PYM binding, thus indicating that the ADP control of the carrier transition was gradually lost. The ADP control could be fully restored through the addition of CL, provided that the phospholipase incubation had been terminated after approximately 1 h. These results will be discussed in relation to an earlier report of tight cardiolipin binding [Beyer, K., & Klingenberg, M. (1985) Biochemistry 24, 3821-3826] and to current structural models of the ADP/ATP carrier protein.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cardiolipins / pharmacology*
  • Carrier Proteins / metabolism*
  • Cattle
  • Cysteine / metabolism
  • Ethylmaleimide / pharmacology
  • Fluorescence
  • Fluorescent Dyes / metabolism
  • Kinetics
  • Magnetic Resonance Spectroscopy
  • Maleimides / metabolism
  • Maleimides / pharmacology
  • Mitochondria, Heart / chemistry*
  • Phospholipases A / metabolism
  • Phospholipases A / pharmacology
  • Phospholipases A2
  • Phospholipids / chemistry
  • Phospholipids / pharmacology
  • Protein Binding / drug effects
  • Protein Conformation
  • Receptors, Cytoplasmic and Nuclear / metabolism*
  • Sulfhydryl Compounds / metabolism
  • Sulfhydryl Reagents / pharmacology

Substances

  • ADP-ATP carrier receptor
  • Cardiolipins
  • Carrier Proteins
  • Fluorescent Dyes
  • Maleimides
  • Phospholipids
  • Receptors, Cytoplasmic and Nuclear
  • Sulfhydryl Compounds
  • Sulfhydryl Reagents
  • N-(3-pyrene)maleimide
  • Phospholipases A
  • Phospholipases A2
  • Cysteine
  • Ethylmaleimide