ADP‐ and oligomycin‐sensitive redox behavior of F₀ b thiol in ATPsynthase depends on neighbored primary structure: Investigations using 14‐C‐labeled alpha lipoic acid

Abstract

Purified ATPsynthase of bovine heart mitochondria has been analyzed for its mobility and reactivity of oligomycin-sensitive sulfhydryl regions in presence of the substrate ADP and oligomycin. Labeling of thiol groups at the hydrophobic F_0 region of the ATPsynthase was increased in the enzyme initially treated with SDS, N-ethylmaleimide and dithiothreitol (modified enzyme). After dialysis or gel permeation the ATPsynthase was treated with [14C] alpha lipoic acid at a molar ratio of 35-85/1 (lipoic acid/ATPsynthase) corresponding to 4-8.6 nmol/mg protein. Under these conditions, ATPase activity of the native enzyme was significantly decreased. After preincubation with ADP, PAGE of the native, [14C] labeled enzyme revealed an increase of radioactivity at a region of 25 kDa deduced to Cys 197 of subunit b. In the modified enzyme the increase in radioactivity was found at 10 kDa. In this context, the sequence Lys-Cys-Ile around Cys 197 of subunit b suggests excessive reactivity of this thiol, as well as ready reversibility by -SH-S-S- interchange. Therefore, previously observed reaction by thiol reagents and antioxidants from outside the mitochondrion can be interpreted with Cys 197 of F0 b. It accounts for sulfhydryl unmasked by binding of ADP at F1.

Protection From Hepatic Ischemia/Reperfusion Injury and Improvement of Liver Regeneration by ??-Lipoic Acid

“…ATP synthesis occurs only if the dithiols between the F1 and the F 0 j part of the enzyme remain functional (27). The lipophilic character of LA allows it to reach the inner membrane of the mitochondrion and to enhance mobility and integrity of the enzyme (28). ATP synthesis may therefore also occur under conditions of ischemia/reperfusion.…”

Different Protection Mechanisms after Pretreatment with Glycine or α-Lipoic Acid in a Rat Model of Warm Hepatic Ischemia

“…The mechanism which could enable LA to influence the prevention of caspase-9 activation, or the release of cytochrome c , remains a matter of speculation. Previous studies have, however, demonstrated that LA interacts directly with an essential enzyme of the respiratory chain, enabling the amino acid to maintain its integrity and function [25] . A further cause for the rise in caspase-3 activity may be an increase in the LPO level in liver tissue in the early phase of IRI [26] .…”

Hepatocellular Injury of Nonischemic Liver Tissue after Selective Clamping in Rats – Protective Action by Pharmacological Pretreatment with Lipoic Acid

“…We recently described similar effects of LA in the IL after ischemia/reperfusion [19] . Moreover, the lipophilic character of LA allows it to reach the membrane of mitochondria [24] . Under these conditions, LA increases production of ATP in vitro.…”