cAMP-dependent tyrosine phosphorylation of subunit I inhibits cytochrome c oxidase activity

J Biol Chem. 2005 Feb 18;280(7):6094-100. doi: 10.1074/jbc.M411335200. Epub 2004 Nov 19.


Signaling pathways targeting mitochondria are poorly understood. We here examine phosphorylation by the cAMP-dependent pathway of subunits of cytochrome c oxidase (COX), the terminal enzyme of the electron transport chain. Using anti-phospho antibodies, we show that cow liver COX subunit I is tyrosinephosphorylated in the presence of theophylline, a phosphodiesterase inhibitor that creates high cAMP levels, but not in its absence. The site of phosphorylation, identified by mass spectrometry, is tyrosine 304 of COX catalytic subunit I. Subunit I phosphorylation leads to a decrease of V(max) and an increase of K(m) for cytochrome c and shifts the reaction kinetics from hyperbolic to sigmoidal such that COX is fully or strongly inhibited up to 10 mum cytochrome c substrate concentrations, even in the presence of allosteric activator ADP. To assess our findings with the isolated enzyme in a physiological context, we tested the starvation signal glucagon on human HepG2 cells and cow liver tissue. Glucagon leads to COX inactivation, an effect also observed after incubation with adenylyl cyclase activator forskolin. Thus, the glucagon receptor/G-protein/cAMP pathway regulates COX activity. At therapeutic concentrations used for asthma relief, theophylline causes lung COX inhibition and decreases cellular ATP levels, suggesting a mechanism for its clinical action.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Allosteric Regulation / drug effects
  • Amino Acid Sequence
  • Animals
  • Cattle
  • Colforsin / pharmacology
  • Crystallography, X-Ray
  • Cyclic AMP / metabolism*
  • Cyclic AMP / pharmacology
  • Cyclic AMP-Dependent Protein Kinases / metabolism*
  • Electron Transport Complex IV / antagonists & inhibitors*
  • Electron Transport Complex IV / chemistry
  • Electron Transport Complex IV / metabolism*
  • Glucagon / pharmacology
  • Kinetics
  • Liver / enzymology
  • Lung / enzymology
  • Models, Molecular
  • Myocardium / enzymology
  • Phosphorylation / drug effects
  • Phosphotyrosine / metabolism*
  • Protein Conformation
  • Protein Subunits / chemistry
  • Protein Subunits / metabolism
  • Theophylline / pharmacology


  • Protein Subunits
  • Colforsin
  • Phosphotyrosine
  • Glucagon
  • Theophylline
  • Cyclic AMP
  • Electron Transport Complex IV
  • Cyclic AMP-Dependent Protein Kinases