Regulation of H2S homeostasis in humans is poorly understood. Therefore, we assessed the importance of individual enzymes in synthesis and catabolism of H2S by studying patients with respective genetic defects. We analyzed sulfur compounds (including bioavailable sulfide) in 37 untreated or insufficiently treated patients with seven ultrarare enzyme deficiencies and compared them to 63 controls. Surprisingly, we observed that patients with severe deficiency in cystathionine β-synthase (CBS) or cystathionine γ-lyase (CSE) - the enzymes primarily responsible for H2S synthesis - exhibited increased and normal levels of bioavailable sulfide, respectively. However, an approximately 21-fold increase of urinary homolanthionine in CBS deficiency strongly suggests that lacking CBS activity is compensated for by an increase in CSE-dependent H2S synthesis from accumulating homocysteine, which suggests a control of H2S homeostasis in vivo. In deficiency of sulfide:quinone oxidoreductase - the first enzyme in mitochondrial H2S oxidation - we found normal H2S concentrations in a symptomatic patient and his asymptomatic sibling, and elevated levels in an asymptomatic sibling, challenging the requirement for this enzyme in catabolizing H2S under physiological conditions. Patients with ethylmalonic encephalopathy and sulfite oxidase/molybdenum cofactor deficiencies exhibited massive accumulation of thiosulfate and sulfite with formation of large amounts of S-sulfocysteine and S-sulfohomocysteine, increased renal losses of sulfur compounds and concomitant strong reduction in plasma total cysteine. Our results demonstrate the value of a comprehensive assessment of sulfur compounds in severe disorders of homocysteine/cysteine metabolism and provide evidence for redundancy and compensatory mechanisms in the maintenance of H2S homeostasis.

Center for Inborn Errors of Metabolism Pediatric Intensive Care Unit University Hospital of Nantes Nantes France

Department of Analytical Chemistry Faculty of Science Charles University Prague Czech Republic

Department of Inborn Errors of Metabolism and Pediatrics The Institute of Mother and Child Warsaw Poland

Department of Molecular Immunology and Toxicology and the National Tumor Biology Laboratory National Institute of Oncology Budapest Hungary

Department of Molecular Immunology and Toxicology and the National Tumor Biology Laboratory National Institute of Oncology Budapest Hungary; Department of Anatomy and Histology ELKH ÁTE Laboratory of Redox Biology University of Veterinary Medicine Budapest Hungary; Chemistry Institute University of Debrecen Debrecen Hungary

Department of Pediatrics and Inherited Metabolic Disorders Charles University 1st Faculty of Medicine and General University Hospital Prague Prague Czech Republic

Department of Pediatrics and Inherited Metabolic Disorders Charles University 1st Faculty of Medicine and General University Hospital Prague Prague Czech Republic; Institute of Rheumatology Prague Czech Republic

Department of Pediatrics Metabolic diseases AOU Città della Salute e della Scienza University of Torino Torino Italy

Department of Pediatrics University Medical Centre Eppendorf Hamburg Germany

Division of Metabolism Bambino Gesù Children's Hospital IRCCS Rome Italy

Genetics and Genomic Medicine Department UCL GOS Institute of Child Health London UK

Inborn Errors of Metabolism Unit Institut de Recerca Sant Joan de Déu and CIBERER ISCIII Barcelona Spain

Institute of Biochemistry Department of Chemistry University of Cologne Cologne Germany

Institute of Medical Genetics and Applied Genomics University of Tübingen Tübingen Germany

Institute of Medical Genetics and Applied Genomics University of Tübingen Tübingen Germany; Department of Neuropediatrics Developmental Neurology and Social Pediatrics University of Tübingen Tübingen Germany

Kinder und Jugendklinik Universitätsklinikum Erlangen Erlangen Germany

Manchester Centre for Genomic Medicine St Mary's Hospital Manchester University NHS Foundation Trust Health Innovation Manchester Manchester United Kingdom

Metabolic Rare Diseases Unit Department of Pediatrics Fondazione MBBM San Gerardo Hospital Monza Italy

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