期刊
PHARMACOLOGICAL RESEARCH
卷 113, 期 -, 页码 116-124出版社
ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD
DOI: 10.1016/j.phrs.2016.08.023
关键词
H2S; Bioenergetics; S-Sulfhydration; Burn; ATP synthase; Cysteine; Mitochondria; Bioenergetics; ATP synthase; Hydrogen sulfide; Burn injury
资金
- Canadian Institutes of Health Research
- Mid-Career Investigator award from the Heart and Stroke Foundation of Ontario
- Discovery Grant from the Natural Sciences and Engineering Research Council of Canada
- Shriners of North America [85800]
- National Institutes of Health [R01 GM107846]
Mammalian cells can utilize hydrogen sulfide (H2S) to support mitochondrial respiration. The aim of our study was to explore the potential role of S-sulfhydration (a H2S-induced posttranslational modification, also known as S-persulfidation) of the mitochondrial inner membrane protein ATP synthase (FIFO ATP synthase/Complex V) in the regulation of mitochondrial bioenergetics. Using a biotin switch assay, we have detected S-sulfhydration of the alpha subunit (ATP5A1) of ATP synthase in response to exposure to H2S in vitro. The H2S generator compound NaHS induced S-sulfhydration of ATP5A1 in HepG2 and HEK293 cell lysates in a concentration-dependent manner (50-300 mu M). The activity of immunocaptured mitochondrial ATP synthase enzyme isolated from HepG2 and HEK293 cells was stimulated by NaHS at low concentrations (10-100 nM). Site-directed mutagenesis of ATP5A1 in HEK293 cells demonstrated that cysteine residues at positions 244 and 294 are subject to S-sulfhydration. The double mutant ATP synthase protein (C244S/C294S) showed a significantly reduced enzyme activity compared to control and the single-cysteine-mutated recombinant proteins (C244S or C294S). To determine whether endogenous H2S plays a role in the basal S-sulfhydration of ATP synthase in vivo, we compared liver tissues harvested from wild-type mice and mice deficient in cystathionine-gamma-lyase (CSE, one of the three principal mammalian H2S-producing enzymes). Significantly reduced S-sulfhydration of ATP5A1 was observed in liver homogenates of CSE-/- mice, compared to wild-type mice, suggesting a physiological role for CSE-derived endogenous H2S production in the S-sulfhydration of ATP synthase. Various forms of critical illness (including burn injury) upregulate H2S-producing enzymes and stimulate H2S biosynthesis. In liver tissues collected from mice subjected to burn injury, we detected an increased S-sulfhydration of ATP5A1 at the early time points post-burn. At later time points (when systemic H2S levels decrease) S-sulfhydration of ATP5A1 decreased as well. In conclusion, H2S induces S-sulfhydration of ATP5A1 at C244 and C294. This post-translational modification may be a physiological mechanism to maintain ATP synthase in a physiologically activated state, thereby supporting mitochondrial bioenergetics. The sulfhydration of ATP synthase may be a dynamic process, which may be regulated by endogenous H2S levels under various pathophysiological conditions. (C) 2016 Elsevier Ltd. All rights reserved.
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