Carbon-Carbon Ligases ; deficiency ; Child ; Gas Chromatography-Mass Spectrometry ; Humans ; Male ; Prognosis

OBJECTIVETo study the activity of vitamin K-dependent gamma-glutamyl carboxylase in patients with calcium oxalate (CaOx) urolithiasis compared with healthy individuals and to assess its relationship to the renal calcium oxalate urolithiasis.

METHODSRenal parenchymas were harvested from urolithic patients and renal tumor patients undergoing nephrectomy. The renal carboxylase activity was evaluated as the radioactivity of [(14)C] labeled sodium bicarbonate in carboxylic reactions in vitro using beta-liquid scintillation counting.

RESULTSSignificantly reduced activity of renal vitamin K-dependent gamma-glutamyl carboxylase was observed in the urolithic group as compared with normal controls (P < 0.01).

CONCLUSIONIt suggests that the reduced carboxylase activity observed in the urolithic patients may play an important role in the course of renal calcium oxalate urolithiasis.

Adult ; Aged ; Calcium Oxalate ; metabolism ; Carbon-Carbon Ligases ; metabolism ; Humans ; Kidney ; enzymology ; Kidney Calculi ; enzymology ; Middle Aged

OBJECTIVE: To explore the genetic basis for a child with clinically suspected 3-methylcrotonyl-coenzyme A carboxylase deficiency (MCCD). METHODS: Genomic DNA was extracted from peripheral blood samples of the proband and her parents. Whole exome sequencing was used to screen pathogenic variant in the proband. Suspected variant was verified by Sanger sequencing. Impact of the variant on the structure and function of protein product was analyzed by using bioinformatic software. RESULTS: Sanger sequencing showed that the proband has carried homozygous missense c.1342G>A (p.Gly448Ala) variant of the MCCC2 gene, for which her mother was a heterozygous carrier. The same variant was not detected in her father. The variant was predicted to be pathogenic by PolyPhen-2 and Mutation Taster software, and the site was highly conserved among various species. Based on the American College of Medical Genetics and Genomics standards and guidelines, the c.1342G>A (p.Gly448Ala) variant of MCCC2 gene was predicted to be likely pathogenic(PM2+PP2-PP5). CONCLUSION The homozygous missense variant of the MCCC2 gene c.1342G>A (p.Gly448Ala) probably underlay the molecular pathogenesis of the proband. Genetic testing has confirmed the clinical diagnosis.
Carbon-Carbon Ligases/genetics* ; Child ; Female ; Humans ; Male ; Mutation, Missense/genetics* ; Pedigree ; Urea Cycle Disorders, Inborn/genetics*

Recombinant strains expressing enzymes for ATP regeneration and L-theanine production were constructed and used for the synthesis of L-theanine. The ppk gene encoding polyphosphate kinase (PPK) from Rhodobacter sphaeroides and gmas gene encoding γ-glutamylmethylamide synthetase (GMAS) from Methylovorus mays were synthesized, and two recombinant plasmids, pETDuet-ppk+gmas and pET21a-ppk+gmas were constructed for co-expression of PPK and GMAS in Escherichia coli BL21(DE3). SDS-PAGE analysis showed that PPK and GMAS were overexpressed in soluble form in both recombinant strains. GMAS-PPK obtained from the recombinant strain containing pET21a-ppk+gmas was more efficient to synthesize L-theanine. After 24 h at 37 ℃ and pH at 7.0, 86.0% yield of L-theanine was achieved with catalytic amount of ATP. This study extends the application of enzymatic ATP regeneration system. In addition, it provides an efficient method for the biosynthesis of L-theanine.
Carbon-Nitrogen Ligases ; genetics ; Electrophoresis, Polyacrylamide Gel ; Escherichia coli ; genetics ; Glutamates ; biosynthesis ; Ligases ; Phosphotransferases (Phosphate Group Acceptor) ; genetics

Pseudoxanthoma elasticum is a multiorgan disorder, characterized by ectopic mineralization of elastic fibers of skin, eyes, and cardiovascular system. The dermatologic manifestations include laxity of skin, as well as cutis laxa-like wrinkling especially on both axillae. The classic forms of pseudoxanthoma elasticum are due to mutations in the ATP-binding cassette subfamily C member 6 (ABCC6) gene, a presumed transmembrane transporter expressed primarily in the liver and the kidneys. Recent case reports of mutations of gamma-glutamyl carboxylase (GGCX) gene describe associations with vitamin K-dependent coagulation factor deficiency and pseudoxanthoma elasticum as well. A 23-year old woman presented with laxity on both axillae and trunk and a history of vitamin K-dependent coagulation factor deficiency, diagnosed 2 years ago. Fragmented and markedly degenerated elastic fibers with calcium deposition were identified on biopsy specimens. Herein, we report a case of autosomal recessive pseudoxanthoma elasticum combined with vitamin K-dependent coagulation factor deficiency.
Axilla ; Biopsy ; Blood Coagulation Factors ; Calcium ; Carbon-Carbon Ligases ; Cardiovascular System ; Elastic Tissue ; Eye ; Female ; Genes, vif ; Humans ; Kidney ; Liver ; Pseudoxanthoma Elasticum ; Skin ; Vitamins

OBJECTIVE: To investigate the genetic characterization of 3-hydroxyisovalerylcarnitine (C5-OH) metabolic abnormality in neonates. METHODS: Fifty two newborns with increased C5-OH, C5-OH/C3 and C5-OH/C8 detected by tandem mass spectrometry during neonatal screening were enrolled in the study. Genomic DNA was extracted from the whole blood samples of 52 cases and their parents. Seventy-nine genes associated with genetic and metabolic diseases including , were targeted by liquid capture technique. Variation information of these genes was examined by high-throughput sequencing and bioinformatic analysis, and then was classified based on the American College of Medical Genetics and Genomics (ACMG) standards and guidelines. The genetic types were classified as wild-type, -maternal-mutation, -paternal-mutation and -mutation. Wilcoxon rank-sum test was performed for the increased multiples of C5-OH calculated in neonatal screening. RESULTS: Twenty one variants (14 novel) were identified in 37 cases, 6 variants (5 novel) in 4 cases. The increased multiple of C5-OH calculated in -maternal-mutation and -mutation groups were significantly higher than that in wild-type group (all <0.05), while there was no significant difference between MCCC1-paternal-mutation group and wild-type group (>0.05). CONCLUSIONS Mutations on and genes are the major genetic causes for the increased C5-OH in neonates, and maternal single heterozygous mutation can contribute to the moderately to severely increased C5-OH.
Carbon-Carbon Ligases ; genetics ; Carnitine ; analogs & derivatives ; metabolism ; Female ; Genetic Testing ; Genetic Variation ; Humans ; Infant, Newborn ; Male ; Mutation ; Neonatal Screening ; Urea Cycle Disorders, Inborn ; genetics

PURPOSE: Seizure associated with fever may indicate the presence of underlying inherited metabolic diseases. The present study was performed to investigate the presence of underlying metabolic diseases in patients with complex febrile seizures, using analyses of urine organic acids. METHODS: We retrospectively analyzed and compared the results of urine organic acid analysis with routine laboratory findings in 278 patients referred for complex febrile seizure. RESULTS: Of 278 patients, 132 had no abnormal laboratory findings, and 146 patients had at least one of the following abnormal laboratory findings: acidosis (n=58), hyperammonemia (n=55), hypoglycemia (n=21), ketosis (n=12). Twenty-six (19.7%) of the 132 patients with no abnormal findings and 104 (71.2%) of the 146 patients with statistically significant abnormalities showed abnormalities on the organic acid analysis (P<0.05). Mitochondrial respiratory chain disorders (n=23) were the most common diseases found in the normal routine laboratory group, followed by PDH deficiency (n=2 ) and ketolytic defect (n=1). In the abnormal routine laboratory group, mitochondrial respiratory chain disorder (n=29) was the most common disease, followed by ketolytic defects (n=27), PDH deficiency (n=9), glutaric aciduria type II (n=9), 3-methylglutaconic aciduria type III (n=6), biotinidase deficiency (n=5), propionic acidemia (n=4), methylmalonic acidemia (n=2), 3-hydroxyisobutyric aciduria (n=2), orotic aciduria (n=2), fatty acid oxidation disorders (n=2), 2-methylbranched chain acyl CoA dehydrogenase deficiency (n=2), 3-methylglutaconic aciduria type I (n=1), maple syrup urine disease (n=1), isovaleric acidemia (n=1), HMG-CoA lyase deficiency (n=1), L-2-hydroxyglutaric aciduria (n=1), and pyruvate carboxylase deficiency (n=1). CONCLUSION: These findings suggest that urine organic acid analysis should be performed in all patients with complex febrile seizure and other risk factors for early detection of inherited metabolic diseases.
Acetyl-CoA C-Acetyltransferase ; Acidosis ; Acyl-CoA Dehydrogenase ; Amino Acid Metabolism, Inborn Errors ; Biotinidase Deficiency ; Brain Diseases, Metabolic, Inborn ; Electron Transport ; Fever ; Humans ; Hydroxybutyrates ; Hyperammonemia ; Hypoglycemia ; Isovaleryl-CoA Dehydrogenase ; Ketosis ; Maple Syrup Urine Disease ; Metabolic Diseases ; Multiple Acyl Coenzyme A Dehydrogenase Deficiency ; Propionic Acidemia ; Pyruvate Carboxylase Deficiency Disease ; Pyruvate Dehydrogenase Complex Deficiency Disease ; Retrospective Studies ; Risk Factors ; Seizures ; Seizures, Febrile

OBJECTIVETo explore the molecular mechanism for a boy suspected with 3-methylcrotonyl-CoA carboxylase deficiency by neonatal screening.

METHODSPCR and Sanger sequencing were used to identify potential mutations of MCCC1 and MCCC2 genes. SIFT and Polyphen-2 software was used to predict the effect of variant on the protein function and conservation of the variant across various species. Human Splicing Finder and Swiss-PdbViewer4.1.0 were applied to analyze the possible mechanism of the variant.

RESULTSFor the proband, a compound heterozygous mutation was discovered in the MCCC1 gene, namely c.539G>T (p.G180V) and c.704_711del (p.A235Vfs*4), which were inherited from his father and mother, respectively. The two mutations have disrupted the protein conformation, which in turn may impact the function of MCC protein.

CONCLUSIONThe compound heterozygous mutations of the MCCC1 gene may contribute to the 3-methylcrotonyl-CoA carboxylase deficiency manifested by the patient.

Amino Acid Sequence ; Base Sequence ; Carbon-Carbon Ligases ; chemistry ; deficiency ; genetics ; DNA Mutational Analysis ; Heterozygote ; Humans ; Infant, Newborn ; Male ; Models, Molecular ; Mutation ; Neonatal Screening ; methods ; Protein Conformation ; Sequence Homology, Amino Acid ; Urea Cycle Disorders, Inborn ; diagnosis ; genetics

Biotin is an important micronutrient that serves as an essential enzyme cofactor. Bacteria obtain biotin either through de novo synthesis or by active uptake from exogenous sources. Mycobacteria are unusual amongst bacteria in that their primary source of biotin is through de novo synthesis. Here we review the importance of biotin biosynthesis in the lifecycle of Mycobacteria. Genetic screens designed to identify key metabolic processes have highlighted a role for the biotin biosynthesis in bacilli growth, infection and survival during the latency phase. These studies help to establish the biotin biosynthetic pathway as a potential drug target for new anti-tuberculosis agents.
Biotin ; biosynthesis ; Carbon-Carbon Ligases ; metabolism ; Carrier Proteins ; metabolism ; Cell Membrane ; metabolism ; Coenzymes ; metabolism ; Fatty Acids ; biosynthesis ; Genes, Bacterial ; Genome, Bacterial ; Metabolic Networks and Pathways ; Molecular Structure ; Mycobacterium Infections ; microbiology ; Mycobacterium tuberculosis ; genetics ; metabolism ; pathogenicity ; physiology ; Virulence

OBJECTIVETo report the clinical diagnosis, treatment and follow-up of children with holocarboxylase synthetas(HCS) deficiency and explore the gene mutation spectrum of the disease.

METHODSEleven children with HCS deficiency were enrolled. Mass spectrometry analysis and biotinidase activity determination were used for diagnosis of HCS deficiency. HCS gene mutations were analyzed by PCR directed sequencing methods. Ten patients received oral biotin treatment (10-40 mg/d). Clinical effects of biotin treatment were observed.

RESULTSAll 11 cases developed apathetic, lethargy and metabolic acidosis at different degrees, and 10 cases presented with skin lesions. The average blood 3-hydroxyisovaleryl-carnitine concentrations and urinary 3-methylcrontonylglycine and methylcitrate concentrations increased significantly. The biotinidase activity increased, being higher over 30% of the normal reference value. Four mutations in HCS gene were identified, and they were c.1522C>T (R508W), c.1088T>A (V363D), c.126G>T (E42D) and c.1994G>C (R665P) (a new variant) and the frequency was 50%, 29%, 7% and 14% respectively. The symptoms disappeared in 10 cases 1-2 weeks after biotin treatment, and blood and urinary abnormal metabolites were gradually reduced to normal 2-6 months after treatment.

CONCLUSIONSHCS deficiency is characterized by nervous system damage, skin lesions and metabolic acidosis. Mass spectrometry analysis, biotinidase activity determination and gene mutation analysis may be helpful in the definite diagnosis of this disorder. The effect of early biotin treatment is satisfactory. The mutations R508W and V363D might be hot-spots in Chinese children with HCS deficiency.

Biotin ; therapeutic use ; Biotinidase ; metabolism ; Carbon-Nitrogen Ligases ; genetics ; Child, Preschool ; Female ; Holocarboxylase Synthetase Deficiency ; diagnosis ; therapy ; Humans ; Infant ; Infant, Newborn ; Male ; Mutation