3-ketoacid CoA Transferase Research Articles

Suppressive Effects of β-Hydroxybutyrate Administration on Lipopolysaccharide-Induced Inflammation in Broiler Chickens.

This study aimed to evaluate the suppressive effects of β-hydroxybutyrate (BHB) administration on lipopolysaccharide (LPS)-induced inflammation in broiler chickens. Twenty-day-old male broiler chickens were randomly allocated to three groups, each of which was treated with saline (control), intraperitoneal administration of LPS [1.5 mg/kg body weight (BW), Escherichia coli O127:B8], or LPS plus BHB (3 mmol/kg BW). Plasma albumin and total protein concentration were significantly reduced by LPS administration, while BHB co-treatment partially attenuated the effects. The LPS treatment significantly induced plasma aspartate and alanine aminotransferase activities, and interleukin (IL)-6 concentration, with the increases suppressed by BHB co-treatment (p < 0.05). The LPS treatment significantly increased the gene expression levels of IL-1β, IL-6, and IL-18 in the spleen and peripheral blood monocytes (PBMC), while the increases were partially attenuated by BHB in the spleen. Relatively higher levels of BHB dehydrogenase 1 and succinyl-CoA:3-ketoacid CoA transferase were observed in the spleen and skeletal muscle, while these gene levels were lower in PBMC and the liver. The present results suggest that BHB can suppress LPS-induced inflammation, in which ketolytic enzyme expression levels may be involved in broiler chickens.

Ketone ester administration improves glycemia in obese mice.

During periods of prolonged fasting/starvation, the liver generates ketones [i.e., β-hydroxybutyrate (βOHB)] that primarily serve as alternative substrates for ATP production. Previous studies have demonstrated that elevations in skeletal muscle ketone oxidation contribute to obesity-related hyperglycemia, whereas inhibition of succinyl CoA:3-ketoacid CoA transferase (SCOT), the rate-limiting enzyme of ketone oxidation, can alleviate obesity-related hyperglycemia. As circulating ketone levels are a key determinant of ketone oxidation rates, we tested the hypothesis that increases in circulating ketone levels would worsen glucose homeostasis secondary to increases in muscle ketone oxidation. Accordingly, male C57BL/6J mice were subjected to high-fat diet-induced obesity, whereas their lean counterparts received a standard chow diet. Lean and obese mice were orally administered either a ketone ester (KE) or placebo, followed by a glucose tolerance test. In tandem, we conducted isolated islet perifusion experiments to quantify insulin secretion in response to ketones. We observed that exogenous KE administration robustly increases circulating βOHB levels, which was associated with an improvement in glucose tolerance only in obese mice. These observations were independent of muscle ketone oxidation, as they were replicated in mice with a skeletal muscle-specific SCOT deficiency. Furthermore, the R-isomer of βOHB produced greater increases in perifusion insulin levels versus the S-isomer in isolated islets from obese mice. Taken together, acute elevations in circulating ketones promote glucose-lowering in obesity. Given that only the R-isomer of βOHB is oxidized, further studies are warranted to delineate the precise role of β-cell ketone oxidation in regulating insulin secretion.NEW & NOTEWORTHY It has been demonstrated that increased skeletal muscle ketone metabolism contributes to obesity-related hyperglycemia. Since increases in ketone supply are key determinants of organ ketone oxidation rates, we determined whether acute elevations in circulating ketones following administration of an oral ketone ester may worsen glucose homeostasis in lean or obese mice. Our work demonstrates the opposite, as acute elevations in circulating ketones improved glucose tolerance in obese mice.

A Novel Mutation in the OXCT1 Gene Causing Succinyl-CoA:3-Ketoacid CoA Transferase (SCOT) Deficiency Starting with Neurologic Manifestations.

Succinyl-CoA:3-oxoacid CoA-transferase (SCOT) deficiency is an inborn error of ketone body utilization characterized by intermittent ketoacidosis crises. This study reports the first Iranian patient with SCOT deficiency who presented with seizure and hypotonia at birth. Accordingly, she was consequently re-hospitalized due to hypotonia and respiratory distress. Laboratory tests revealed hyperammonemia, ketonuria, and metabolic acidosis. Besides, the plasma glucose level was normal without any other abnormality. Despite treatment with high-dose bicarbonate, severe acidosis persisted. Poor response to treatment raised a significant diagnostic challenge among specialists until genetic investigation identified a homozygous nonsense mutation (c.79G>T; p.Gly27*) in the OXCT1 gene (NM_000436), causing SCOT deficiency. Genetic studies help clinicians achieve a definite diagnosis of such metabolic disorders. In this case, the accurate and early diagnosis of SCOT deficiency opened new therapeutic possibilities, including frequent carbohydrate-rich meals and low fat and protein diet. Moreover, our findings expand the mutational and clinical spectrum of SCOT deficiency.

The Antipsychotic Dopamine 2 Receptor Antagonist Diphenylbutylpiperidines Improve Glycemia in Experimental Obesity by Inhibiting Succinyl-CoA:3-Ketoacid CoA Transferase.

Despite significant progress in understanding the pathogenesis of type 2 diabetes (T2D), the condition remains difficult to manage. Hence, new therapeutic options targeting unique mechanisms of action are required. We have previously observed that elevated skeletal muscle succinyl CoA:3-ketoacid CoA transferase (SCOT) activity, the rate-limiting enzyme of ketone oxidation, contributes to the hyperglycemia characterizing obesity and T2D. Moreover, we identified that the typical antipsychotic agent pimozide is a SCOT inhibitor that can alleviate obesity-induced hyperglycemia. We now extend those observations here, using computer-assisted in silico modeling and invivo pharmacology studies that highlight SCOT as a noncanonical target shared among the diphenylbutylpiperidine (DPBP) drug class, which includes penfluridol and fluspirilene. All three DPBPs tested (pimozide, penfluridol, and fluspirilene) improved glycemia in obese mice. While the canonical target of the DPBPs is the dopamine 2 receptor, studies in obese mice demonstrated that acute or chronic treatment with a structurally unrelated antipsychotic dopamine 2 receptor antagonist, lurasidone, was devoid of glucose-lowering actions. We further observed that the DPBPs improved glycemia in a SCOT-dependent manner in skeletal muscle, suggesting that this older class of antipsychotic agents may have utility in being repurposed for the treatment of T2D.

118-LB: The Antipsychotic D2 Receptor Antagonist Diphenylbutylpiperidines Improve Glycemia in Experimental Obesity by Inhibiting Succinyl-CoA—3-Ketoacid CoA transferase

Purpose: Despite significant progress in understanding the pathogenesis of type 2 diabetes (T2D) , it remains difficult to manage, hence, new therapeutic options are required. We previously observed that elevated skeletal muscle succinyl CoA:3-ketoacid CoA transferase (SCOT) activity, the rate-limiting enzyme of ketone (KB) oxidation, contributes to obesity-induced hyperglycemia. Moreover, we identified that the antipsychotic agent, pimozide, is a SCOT inhibitor with glucose-lowering actions. In silico molecular modeling determined that a wide range of DPBPs can theoretically inhibit SCOT activity, therefore, we determined whether the DPBP drug class could be repurposed for the treatment of T2D. Methods: 8-week-old male wild-type and muscle-specific/brain-specific SCOT knockout (SCOTMuscleKO, SCOTBrainKO) mice were subjected to experimental obesity via consumption of a high fat, high sugar diet for 12-weeks. Lean control mice received a low fat, low sugar diet. At 8-weeks, lean and obese mice were treated with DPBPs (penfluridol, fluspirilene, pimozide (10mg/kg)) once every 2 days via oral gavage for 14-days, following which circulating KB levels and glucose homeostasis were assessed. To rule out a contribution of the canonical actions of DPBPs as dopamine 2 (D2) receptor antagonists to DPBP-mediated glucose-lowering, obese mice were treated with the structurally unrelated D2 receptor antagonist, lurasidone (10 mg/kg) . Results: All tested DPBPs improved glucose homeostasis in obese mice through a mechanism dependent on the inhibition of both brain and muscle SCOT activity. Treatment with lurasidone failed to improve glycemia in obese mice, thus consistent with a SCOT-dependent mechanism of action. Conclusions: Our findings suggest all DPBPs have glucose-lowering actions and therefore may have clinical utility in being repurposed for the treatment of T2D. Disclosure S. Tabatabaei dakhili: None. C. A. Velazquez: None. P. A. Crawford: Advisory Panel; Abbott Diabetes, Johnson &amp; Johnson Global Services. M. Glover: None. R. Al batran: None. J. R. Ussher: None. R. Abou farraj: None. A. A. Greenwell: None. C. T. Saed: None. K. Yang: None. K. Gopal: None. J. S. F. Chan: None. C. Lee: None. F. Eaton: None. Funding Canadian Institutes of HealthResearch

Ketones Improve the Mitochondrial Coupling Efficiency of Brown Adipocytes

Brown adipose tissue (BAT) thermogenesis is an energy‐demanding response, and this capacity can be leveraged to dissipate excess energy in obesity and its associated metabolic complications. Therefore, understanding the regulatory mechanisms of BAT mitochondrial uncoupling and thermogenesis is crucial. Ketone bodies are endogenous metabolites mainly found during starvation and carbohydrate restriction; however, their regulatory role on BAT thermogenesis is elusive. In this study, we aim to investigate the potential metabolic reprogramming induced by ketones on BAT. We hypothesize that ketones are alternative fuels for BAT during energy/carbohydrate shortage and decrease mitochondrial uncoupling to counterbalance the systemic energy deficit. In mice, ketogenic diet reduces core body temperature by 0.4˚C. Using Seahorse respirometry in differentiated brown adipocytes, we found that ketones improve mitochondrial coupling efficiency (ATP‐linked respiration/ basal respiration) by 30%, implying an attempt to decrease thermogenesis. The gene Oxct1, encoding succinyl‐CoA:3‐ketoacid CoA transferase that is required for ketolysis, is abundantly expressed by brown adipocytes. BAT‐specific Oxct1 knockout renders mice more resistant to cold‐induced decrease in core body temperature during fasting or ketogenic diet, via mechanisms independent of UCP1. Together, our data suggest that ketones potentially communicate hunger to brown adipocytes, thus promoting mitochondrial coupling efficiency for energy conservation.

Succinyl-CoA:3-ketoacid CoA transferase (SCOT) deficiency presenting as ketosis with hyperglycemia: A case report

We describe a case of previously normal 22-month-old male who presented with vomiting, loose motions, lethargy, and seizures for 1 day. He had high anion gap metabolic acidosis, hyperglycemia, and ketosis managed elsewhere as diabetic ketoacidosis. Early management included hemodynamic stabilization, mechanical ventilation, correction of metabolic acidosis, seizure control, and later peritoneal dialysis. He was finally diagnosed with succinyl CoA 3 ketoacid transferase deficiency, a disorder of ketone metabolism usually presents with metabolic acidosis, ketosis, and hypoglycemia. Most cases present during the neonatal period (50%) and remain up to 20 months of age. The pathognomic finding is a nonspecific elevation on ketones in urine gas chromatography–mass spectrometry. Genomic sequencing is confirmatory.

Clinical variability and outcome of succinyl-CoA:3-ketoacid CoA transferase deficiency caused by a single OXCT1 mutation: Report of 17 cases.

Succinyl‐CoA:3‐ketoacid CoA transferase (SCOT) deficiency is an inherited metabolic disease caused by mutated OXCT1 gene resulting in recurrent ketoacidosis. Analysis of longitudinal data in such an ultra‐rare disease is warranted to delineate genotype–phenotype correlations and management outcome. A retrospective analysis of 17 patients, from nine unrelated families, with SCOT deficiency who were followed up in the Medical Genetics Clinic at King Faisal Specialist Hospital and Research Centre was conducted. All the patients were homozygous for p.R468C in OXCT1 gene. Most of the patients (n = 15, 88.2%) were symptomatic presenting with recurrent ketoacidosis, the onset of which ranged from 6 months to 4 years (median 2 years). A striking inter‐ and intrafamilial variability that ranged from being entirely asymptomatic to death during the first episode. All patients were instructed to avoid fasting, restrict protein in diet, and receive carnitine supplementation. However, there was no correlation between following instructions of chronic management and outcome. Most of the patients had their crises resolved and all of them had normal neurodevelopmental outcome. Our data suggest that SCOT deficiency caused by homozygous p.R468C has variable clinical presentation and incomplete penetrance. The apparent lack of correlation between protein restriction +/− carnitine supplementation and outcome suggests that chronic dietary restriction may not be warranted. However, a longer follow‐up on larger and heterogenous cohort of cases is needed before a clear conclusion on the long‐term management can be reached.

Succinyl-CoA: 3-Ketoacid CoA-Transferase Deficiency in a Saudi Girl

Objective: Succinyl-CoA:3-ketoacid CoA transferase (SCOT) deficiency is a rare genetic disorder of ketone utilization and isoleucine catabolism caused by mutations in the OXCT1 gene,&#x0D; Case: A Saudi girl case of SCOT deficiency confirmed by genetic analysis has been reported in this study. A 5-year-old girl presented to the emergency with the first episode of severe metabolic ketoacidosis after a febrile illness. On admission, she was drowsy lethargic, and severely dehydrated needs to admit in a highly dependent area. Initial investigations were done during the crisis showed refractory severe metabolic acidosis (pH of 7.18, HCO3- of 7.4 mmol/L), normal ammonia, lactic acidosis, and urine organic acid profile revealed elevations in 3-hydroxybutyrate and acetoacetate. Genetic analysis was done by CentoMito Comprehensive (Large extended screening panel), sequencing of OXCT1 gene revealed that the proband is homozygous for the missense likely pathogenic variant c.1402C&gt;T p.(Arg468Cys) confirming the diagnosis of SCOT deficiency.&#x0D; Conclusion: This is the first Saudi child with succinyl-CoA:3-ketoacid CoA transferase (SCOT) deficiency case report as searched in the literature. This case highlights the importance of suspecting SCOT deficiency in the differential diagnosis of pediatric metabolic ketoacidosis in preventing life-threatening of severe Metabolic ketoacidosis

Beneficial effects on kidney during treatment with sodium-glucose cotransporter 2 inhibitors: proposed role of ketone utilization.

Modestly elevated circulating levels of the ketone β-hydroxybutyrate (βOHB) during treatment with sodium-glucose cotransporter 2 (SGLT2) inhibitors cause different beneficial effects on organs and cells, depending on the succinyl-CoA:3-ketoacid CoA transferase (SCOT) level. In the failing heart, SCOT is highly expressed/up-regulated, and thus, βOHB may be an energy source, in addition to fat and glucose oxidation. However, SCOT is not highly expressed/down-regulated in the kidney, and thus, βOHB may cause different beneficial effects, rather than acting as an alternative energy source in patients with chronic kidney disease (CKD). βOHB is an endogenous and specific inhibitor of class I histone deacetylases (HDACs) and the NLRP3 inflammasome, accumulates in the kidney because of its decreased utilization as an energy source due to the down-regulation of SCOT, and may induce beneficial effects such as inhibiting inflammation, oxidative stress, and fibrosis. In addition to restoring tubulo-glomerular feedback and improving renal proximal tubule oxygenation, SGLT2 inhibitors may play a renoprotective role by way of βOHB in patients with CKD.

Insulin resistance and heart failure during treatment with sodium glucose cotransporter 2 inhibitors: proposed role of ketone utilization

Sodium glucose cotransporter 2 (SGLT2) inhibitors reduce the rate of hospitalization for heart failure in individuals with type 2 diabetes, but the underlying mechanisms remain elusive. Modestly elevated circulating β-hydroxybutyrate (βOHB) during treatment with SGLT2 inhibitors causes different beneficial effects on organs and cells, depending on succinyl-CoA:3-ketoacid CoA transferase (SCOT) levels. In the heart, in which SCOT is highly expressed/up-regulated, βOHB may be an alternative energy source apart from fat and glucose oxidation. The type 2 diabetic failing heart may be energy inefficient. In skeletal muscle, in which SCOT is not highly expressed/down-regulated, βOHB may cause antioxidant effects, resulting in amelioration of insulin resistance, which could lead to improvement in cardiac insulin resistance with metabolic, endocrine, and cytokine alterations. Although various mechanisms have been suggested, we postulate that the potential impact of SGLT2 inhibitors on heart failure lies in fuel energetics and amelioration of insulin resistance with ketone utilization depending upon SCOT levels.

Identification of a novel OXCT1 frameshift mutation by whole-exome sequencing and evidence for nonsense-mediated mRNA decay

Background: Autosomal recessive disorders are the most common forms of hereditary diseases in consanguineous populations. Despite their frequencies, the diagnosis of these disorders continues to be a challenging task, given their extreme genetic heterogeneities. However, since the discovery of next-generation sequencing-based techniques, the number of pathogenic mutations linked to autosomal recessive disorders has increased dramatically due to the time and cost-effectiveness of these methods. Succinyl-CoA: 3-ketoacid CoA transferase (SCOT) deficiency is considered as one of these rare autosomal recessive genetic disorders. Aim and Objectives: In the present study, we aimed to identify the responsible mutation in a UAE family with autosomal recessive SCOT deficiency. Materials and Methods: Whole-exome sequencing was performed in the affected individual. Sanger sequencing was used to confirm the existence and the segregation of the novel pathogenic mutation. Real-time polymerase chain reaction (PCR) was used to measure the expression of the OXCT1 gene in the mRNA of affected, carrier and normal individuals. Results: Our analysis revealed a new frameshift mutation. Sanger sequencing confirmed its homozygosity in the patient and its cosegregation with the disease in the studied family. Using real-time PCR, we showed that this new frameshift mutation affects the OXCT1-mRNA by nonsense-mediated mRNA decay (NMD). Conclusion: To the best of our knowledge, this is the first study associating SCOT deficiency with an OXCT1 frameshift mutation in the world. In addition, we report for the first time by studying this new OXCT1 mutation evidence for an NMD effect associated with SCOT mutations.

34-OR: Maladaptive Increases in Skeletal Muscle Ketone Body Oxidation Contribute to Dysglycemia in Mice Subjected to Experimental Obesity

Type 2 diabetes (T2D) is a rapidly growing health epidemic, with intense investigation on whether perturbations in skeletal muscle carbohydrate and fatty acid metabolism contribute to obesity-induced insulin resistance and/or T2D. Conversely, whether perturbations in skeletal muscle ketone body oxidation take place in the pathology of obesity-induced insulin resistance/T2D remains unclear. Therefore, our goal was to investigate potential alterations in ketone body metabolism during the pathology of obesity-induced T2D, and to elucidate whether such changes are adaptive or maladaptive. C57BL/6J mice were subjected to low-fat (lean) or high-fat (obese) diet supplementation for 12 weeks. Obese mice exhibited glucose intolerance, which was associated with decreased circulating βOHB and elevated circulating glucose levels. In addition, obese mice demonstrated a marked increase in mRNA and protein expression of succinyl CoA:3-ketoacid CoA transferase (SCOT), the rate limiting enzyme of ketone body oxidation, within gastrocnemius muscles, which was associated with an increase in SCOT enzymatic activity. Of interest, SCOT activity requires succinyl-CoA as a substrate, which is also required for protein succinylation. Accordingly, siRNA knockdown of Oxct1 augmented lysine succinylation, whereas Oxct1 overexpression abolished lysine succinylation. Additionally, Oxct1 knockdown in C2C12 myotubes increased AMPK phosphorylation, which was recapitulated in mice with an Oxct1 skeletal muscle-specific deficiency (Oxct1SkM-/-). Intriguingly, Oxct1SkM-/- mice were protected against obesity-induced dysglycemia and insulin resistance. Our results suggest that ketone body metabolism is augmented in skeletal muscle during the pathology of obesity, which may contribute to dysglycemia and insulin resistance in T2D via potentially modifying protein succinylation and/or AMPK activity. Disclosure R.Al batran: None. K.Gopal: None. J.J.Chahade: None. J.R.Ussher: None.

Report of a case with succinyl-CoA: 3-ketoacid CoA transferase (SCOT)

Report of a case with succinyl-CoA: 3-ketoacid CoA transferase (SCOT)

Inborn errors of ketone body utilization.

Succinyl-CoA:3-ketoacid CoA transferase (SCOT) deficiency and mitochondrial acetoacetyl-CoA thiolase (beta-ketothiolase or T2) deficiency are classified as autosomal recessive disorders of ketone body utilization characterized by intermittent ketoacidosis. Patients with mutations retaining no residual activity on analysis of expression of mutant cDNA are designated as severe genotype, and patients with at least one mutation retaining significant residual activity, as mild genotype. Permanent ketosis is a pathognomonic characteristic of SCOT-deficient patients with severe genotype. Patients with mild genotype, however, may not have permanent ketosis, although they may develop severe ketoacidotic episodes similar to patients with severe genotype. Permanent ketosis has not been reported in T2 deficiency. In T2-deficient patients with severe genotype, biochemical diagnosis is done on urinary organic acid analysis and blood acylcarnitine analysis to observe characteristic findings during both ketoacidosis and non-episodic conditions. In Japan, however, it was found that T2-deficient patients with mild genotype are common, and typical profiles were not identified on these analyses. Based on a clinical study of ketone body utilization disorders both in Japan and worldwide, we have developed guidelines for disease diagnosis and treatment. These diseases are treatable by avoiding fasting and by providing early infusion of glucose, which enable the patients to grow without sequelae.

Structure of succinyl-CoA:3-ketoacid CoA transferase from Drosophila melanogaster.

Succinyl-CoA:3-ketoacid CoA transferase (SCOT) plays a crucial role in ketone-body metabolism. SCOT from Drosophila melanogaster (DmSCOT) was purified and crystallized. The crystal structure of DmSCOT was determined at 2.64 Å resolution and belonged to space group P212121, with unit-cell parameters a=76.638, b=101.921, c=122.457 Å, α=β=γ=90°. Sequence alignment and structural analysis identified DmSCOT as a class I CoA transferase. Compared with Acetobacter aceti succinyl-CoA:acetate CoA transferase, DmSCOT has a different substrate-binding pocket, which may explain the difference in their substrate specificities.

Molecular Basis of Two-Exon Skipping (Exons 12 and 13) by c.1248+5g&gt;a inOXCT1Gene: Study on Intermediates ofOXCT1Transcripts in Fibroblasts

The molecular basis of simultaneous two-exon skipping induced by a splice-site mutation has yet to be completely explained. The splice donor site mutation c.1248+5g>a (IVS13) of the OXCT1 gene resulted predominantly in skipping of exons 12 and 13 in fibroblasts from a patient (GS23) with succinyl-CoA:3-ketoacid CoA transferase (SCOT) deficiency. We compared heteronuclear RNA (hnRNA) intermediates between controls' and GS23's fibroblasts. Our strategy was to use RT-PCR of hnRNA to detect the presence or absence of spliced exon clusters in RNA intermediates (SECRIs) comprising sequential exons. Our initial hypothesis was that a SECRI comprising exons 12 and 13 was formed first followed by skipping of this SECRI in GS23 cells. However, such a pathway was revealed to be not a major one. Hence, we compared the intron removal of SCOT transcript between controls and GS23. In controls, intron 11 was the last intron to be spliced and the removal of intron 12 was also rather slow and occurred after the removal of intron 13 in a major pathway. However, the mutation in GS23 cells resulted in retention of intron 13, thus causing the retention of introns 12 and 11. This "splicing paralysis" may be solved by skipping the whole intron 11-exon 12-intron 12-exon 13-mutated intron 13, resulting in skipping of exons 12 and 13.

A structural mapping of mutations causing succinyl‐CoA:3‐ketoacid CoA transferase (SCOT) deficiency

Succinyl-CoA:3-ketoacid CoA transferase (SCOT) deficiency is a rare inherited metabolic disorder of ketone metabolism, characterized by ketoacidotic episodes and often permanent ketosis. To date there are ∼20 disease-associated alleles on the OXCT1 gene that encodes the mitochondrial enzyme SCOT. SCOT catalyzes the first, rate-limiting step of ketone body utilization in peripheral tissues, by transferring a CoA moiety from succinyl-CoA to form acetoacetyl-CoA, for entry into the tricarboxylic acid cycle for energy production. We have determined the crystal structure of human SCOT, providing a molecular understanding of the reported mutations based on their potential structural effects. An interactive version of this manuscript (which may contain additional mutations appended after acceptance of this manuscript) may be found on the web address: http://www.thesgc.org/jimd/SCOT.Electronic supplementary materialThe online version of this article (doi:10.1007/s10545-013-9589-z) contains supplementary material, which is available to authorized users.

Clinical and molecular characterization of five patients with succinyl-CoA:3-ketoacid CoA transferase (SCOT) deficiency

Succinyl-CoA:3-ketoacid CoA transferase (SCOT) deficiency is an inborn error of ketone body metabolism and causes episodic ketoacidosis. We report clinical and molecular analyses of 5 patients with SCOT deficiency. Patients GS07, GS13, and GS14 are homozygotes of S405P, L327P, and R468C, respectively. GS17 and GS18 are compound heterozygotes for S226N and A215V, and V404F and E273X, respectively. These mutations have not been reported previously. Missense mutations were further characterized by transient expression analysis of mutant cDNAs. Among 6 missense mutations, mutants L327P, R468C, and A215V retained some residual activities and their mutant proteins were detected in immunoblot analysis following expression at 37 °C. They were more stable at 30 °C than 37 °C, indicating their temperature sensitive character. The R468C mutant is a distinct temperature sensitive mutant which retained 12% and 51% of wild-type residual activities at 37 and 30 °C, respectively. The S226N mutant protein was detected but retained no residual activity. Effects of missense mutations were predicted from the tertiary structure of the SCOT molecule. Main effects of these mutations were destabilization of SCOT molecules, and some of them also affected catalytic activity. Among 5 patients, GS07 and GS18 had null mutations in both alleles and the other three patients retained some residual SCOT activities. All 5 developed a first severe ketoacidotic crisis with blood gas pH < 7.1, and experienced multiple ketoacidotic decompensations (two of them had seven such episodes). In general, the outcome was good even following multiple ketoacidotic events. Permanent ketosis or ketonuria is considered a pathognomonic feature of SCOT deficiency. However, this condition depends not only on residual activity but also on environmental factors.

A neonatal‐onset succinyl‐CoA:3‐ketoacid CoA transferase (SCOT)‐deficient patient with T435N and c.658‐666dupAACGTGATT p.N220_I222dup mutations in the OXCT1 gene

Succinyl-CoA:3-ketoacid CoA transferase (SCOT) deficiency causes episodic ketoacidotic crises and no apparent symptoms between them. Here, we report a Japanese case of neonatal-onset SCOT deficiency. The male patient presented a severe ketoacidotic crisis, with blood pH of 7.072 and bicarbonate of 5.8 mmol/L at the age of 2 days and was successfully treated with intravenous infusion of glucose and sodium bicarbonate. He was diagnosed as SCOT deficient by enzymatic assay and mutation analysis. At the age of 7 months, he developed a second ketoacidotic crisis, with blood pH of 7.059, bicarbonate of 5.4 mmol/L, and total ketone bodies of 29.1 mmol/L. He experienced two milder ketoacidotic crises at the ages of 1 year and 7 months and 3 years and 7 months. His urinary ketone bodies usually range from negative to 1+ but sometimes show 3+ (ketostix) without any symptoms. Hence, this patient does not show permanent ketonuria, which is characteristic of typical SCOT-deficient patients. He is a compound heterozygote of c.1304C > A (T435N) and c.658-666dupAACGTGATT p.N220_I222dup. mutations in the OXCT1 gene. The T435N mutation was previously reported as one which retained significant residual activity. The latter novel mutation was revealed to retain no residual activity by transient expression analysis. Both T435N and N220_I222 lie close to the SCOT dimerization interface and are not directly connected to the active site in the tertiary structure of a human SCOT dimer. In transient expression analysis, no apparent interallelic complementation or dominant negative effects were observed. Significant residual activity from the T435N mutant allele may prevent the patient from developing permanent ketonuria.