Journal of Bioenergetics and Biomembranes

, Volume 13, Issue 5–6, pp 393–409 | Cite as

Effect of chemical modifiers of amino acid residues on proton conduction by the H+-ATPase of mitochondria

  • F. Guerrieri
  • S. Papa
Research Papers

Abstract

The effect of chemical modifiers of amino acid residues on the proton conductivity of H+-ATPase in “inside out” submitochondrial particles has been studied. Treatment of submitochondrial particles prepared in the presence of EDTA (ESMP) with the arginine modifiers, phenylglyoxal or butanedione, or the tyrosine modifier, tetranitromethane, caused inhibition of the ATPase activity. Phenylglyoxal and tetranitromethane also caused inhibition of the anaerobic release of respiratory ΔμH+ in ESMP as well as in particles deprived of F1 (USMP). Butanedione treatment caused, on the contrary, acceleration of anaerobic proton release in both particles. The inhibition of proton release caused by phenylglyoxal and tetranitromethane exhibited in USMP a sigmoidal titration curve. The same inhibitory pattern was observed with oligomycin and withN,N′-dicyclohexylcarbodiimide. In ESMP, relaxation of ΔμH+ exhibited two first-order phases, both an expression of the H+ conductivity of the ATPase complex. The rapid phase results from transient enhancement of H+ conduction caused by respiratory ΔμH+ itself. Oligomycin,N,N′-dicyclohexylcarbodiimide, and tetranitromethane inhibited both phases of H+ release, and butanedione accelerated both. Phenylglyoxal inhibited principally the slow phase of H+ conduction. In USMP, H+ release followed simple first-order kinetics. Oligomycin depressed H+ release, enhanced respiratory ΔμH+, and restored the biphasicity of H+ release. Phenylglyoxal and tetranitromethane inhibited H+ release in USMP without modifying its first-order kinetics. Butanedione treatment caused biphasicity of H+ release from USMP, introducing a very rapid phase of H+ release. Addition of soluble F1 to USMP also restored biphasicity of H+ release. A mechanism of proton conduction by F o is discussed based on involvement of tyrosine or other hydroxyl residues, in series with the DCCD-reactive acid residue. There are apparently two functionally different species of arginine or other basic residues: those modified by phenylglyoxal, which facilitate H+ conduction, and those modified by butanedione, which retard H+ diffusion.

Key Words

H+-ATPase proton conduction amino acid modification 

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Copyright information

© Plenum Publishing Corporation 1981

Authors and Affiliations

  • F. Guerrieri
    • 1
  • S. Papa
    • 1
  1. 1.Institute of Biological Chemistry, Faculty of Medicine and Center for the Study of Mitochondria and Energy Metabolism, C.N.R.University of BariBariItaly

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