Analysis of MCCC2 gene variant in a pedigree affected with 3-methylcrotonyl coenzyme A carboxylase deficiency / 中华医学遗传学杂志

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.

Genetic analysis of newborns with abnormal metabolism of 3-hydroxyisovalerylcarnitine / 浙江大学学报·医学版

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).@*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.

A novel compound heterozygous mutation causing 3-methylcrotonyl-CoA carboxylase deficiency / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To explore the molecular mechanism for a boy suspected with 3-methylcrotonyl-CoA carboxylase deficiency by neonatal screening.</p><p><b>METHODS</b>PCR 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.</p><p><b>RESULTS</b>For 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.</p><p><b>CONCLUSION</b>The compound heterozygous mutations of the MCCC1 gene may contribute to the 3-methylcrotonyl-CoA carboxylase deficiency manifested by the patient.</p>

Follow up and gene mutation analysis in cases suspected as 3-methylcrotonyl-coenzyme A carboxylase deficiency by neonatal screening / 中华儿科杂志

<p><b>OBJECTIVE</b>3-Methylcrotonyl-coenzyme A carboxylase deficiency (MCCD) is an autosomal recessive inborn error of leucine catabolism. The cases suspected as MCCD detected by neonatal screening are not rare. The aim of the study was to investigate the clinical outcomes in cases suspected as MCCD by neonatal screening. The second aim was to investigate the mutation spectrum of MCC gene in Chinese population and hotspot mutation.</p><p><b>METHOD</b>Forty-two cases (male 33, female 9) , who had higher blood 3-hydroxy-isovalerylcarnitine (C5-OH) levels(cut-off <0.6 µmol/L) detected by neonatal screening using MS/MS, were recruited to this study during Sept.2011 to Mar.2013. The C5-OH concentrations were [0.84 (0.61-20.15) µmol/L] in 42 cases at the screening recall. Five cases were firstly diagnosed as maternal MCCD, 6 cases as benign MCCD and 31 cases were suspected as MCCD. To follow up the height, weight, mental development, blood C5-OH concentrations and urinary 3-methylcrotonyl-glycine (3-MCG) and 3-hydroxy isovalerate (3-HIVA) in order to investigate the clinical outcome. The MCCC1 and MCCC2 gene mutation were analyzed for some cases. The novel gene variants were evaluated, and the influence of novel missense variants on the protein structure and function were predicted by PolyPhen-2, SIFT, UniProt and PDB software.</p><p><b>RESULT</b>(1) Forty-two cases had no symptoms, their physical and mental development were normal in the last visit at the median ages of 29 months, the oldest age of follow up was nearly 9 years. (2) Gene mutation analysis was performed for 29 cases with informed consent signed by parents.Fourteen different mutations were identified in 19 cases. The mutations in MCCC1 gene accounted for 86%, the most common mutation was c.ins1680A, (accounted for 40%). Nine kinds of novel variant were detected including 211AG>CC/p.Q74P, c.295G>A/p.G99S, c.764A>C/p.H255P, c.964G>A/p. E322K, c.1331G>A/p.R444H, c.1124delT, c.39_58del20, c.1518delG, c.639+2T>A.Other 3 kinds of mutation in MCCC1 gene and 2 kinds of mutation in MCCC2 gene have been reported previously; the amino acid of mutant positions of five kinds of novel missense variant are almost highly conserved. These missense variants were predicted to cause change of human MCC protein side chain structure by changing hydrogen bonding, size of amino acid residue and electric charge, and predicted to damage the protein function possibly according to PolyPhen-2 and PDB analysis. So these novel variants may be disease-causing mutations. No mutation were detected in 10 cases. (3) Blood concentrations of C5-OH when screening, recall and end of follow-up in maternal MCCD was 3.50 (1.63-11.43), 1.84 (1.00-9.30), 0.27 (0.26-5.81) µmol/L. There was a significant downward trend.In contrast, benign MCCD group was 8.20 (3.60-9.60), 9.67 (3.88-20.15), 23.0 (5.87-49.10) µmol/L.It showed a rising trend. Children's urinary 3-MCG of benign MCCD group was found abnormally elevated in 4 cases (100%) when they were recalled.</p><p><b>CONCLUSION</b>A certain number of cases with MCCD or suspected as MCCD in this study had no symptoms and normal physical and mental development after follow-up to oldest age of nearly 9 years. The mutation in MCCC1 gene is common, nine novel mutations were found, c.ins1680A may be a hotspot mutation in Chinese population. The urinary GC/MS analysis and blood MS/MS analysis for mother should be routinely performed for all cases with high blood C5-OH level detected by neonatal screening.</p>

Pseudoxanthoma Elasticum Accompanied with Vitamin K-dependent Coagulation Factor Deficiency / 대한피부과학회지

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.

Clinical and mutational features of maternal 3-methylcrotonyl coenzyme deficiency / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To report on 5 patients with maternal 3-methylcrotonyl coenzyme A carboxylase deficiency (MCCD) and to confirm the clinical diagnosis through mutation analysis.</p><p><b>METHODS</b>Five neonates with higher blood 3-hydroxy isovalerylcarnitine (C5-OH) concentration detected upon newborn screening with tandem mass spectrometry and their mothers were recruited. Urinary organic acids were analyzed with gas chromatography mass spectrometry. Gene mutation and protein function analysis were performed by PCR direct sequencing and PolyPhen-2 software.</p><p><b>RESULTS</b>Higher blood C5-OH concentrations (5.11-21.77 μmol/L) and abnormal 3-hydroxy isovalerate and 3-methylcrotonyl glycine in urine were detected in the five asymptomatic mothers, who were diagnosed as benign MCCD. Higher C5-OH concentration was also detected in their neonates by tandem mass spectrometry, which had gradually decreased to normal levels in three neonates. Four new variations, i.e., c.ins1680A(25%), c.203C > T (p.A68V), c.572T > C (p.L191P) and c.639+5G > T were detected in the MCCC1 gene, in addition with 2 mutations [c.1406G > T (p.R469L, novel variation) and c.592C > T (p.Q198X)]. The novel variations were predicted to have affected protein structure and function.</p><p><b>CONCLUSION</b>For neonates with higher C5-OH concentration detected upon neonatal screening, their mothers should be also tested to rule out MCCD. Mutations in MCCC1 gene are quite common.</p>

Biotin biosynthesis in Mycobacterium tuberculosis: physiology, biochemistry and molecular intervention

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.

Impact of five genetic polymorphisms on inter-individual variation in warfarin maintenance dose / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To investigate the effect of genetic polymorphisms in VKORC1, CYP2C9, GGCX, EPHX1, APOE genes on inter-individual variation in warfarin maintenance dose.</p><p><b>METHODS</b>Two hundred and forty-nine patients with stable warfarin dose were enrolled in this study, and the clinical data and blood samples of the patients were collected. Genotypes for the 5 genes were determined by using PCR and denaturing high performance liquid chromatography (DHPLC) assay. The warfarin maintenance doses were compared among patients with different genotypes of the 5 genes, and a warfarin stable dosing algorithm was derived based on genetic and non-genetic factors.</p><p><b>RESULTS</b>Of the 5 genes, VKORC1, CYP2C9 and GGCX were associated with warfarin stable dose. The multiple linear regression analysis indicated that VKORC1, CYP2C9 and GGCX genes, age and weight, had significant influence on inter-individual variation in warfarin stable dose, which contributed 30.2%, 22.8%, 1.5%, 4.7% and 6.7% respectively. The warfarin stable dosing algorithm acquired from the optimal regression model could explain 57.8% variation in warfarin dose.</p><p><b>CONCLUSION</b>This study suggested that genetic factors are the major determinants of the warfarin maintenance dose, and warfarin stable dosing algorithm may be useful for helping clinicians to prescribe warfarin with greater safety and efficiency.</p>

Decreased renal vitamin K-dependent gamma-glutamyl carboxylase activity in calcium oxalate calculi patients / 中华医学杂志(英文版)

<p><b>OBJECTIVE</b>To 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.</p><p><b>METHODS</b>Renal 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.</p><p><b>RESULTS</b>Significantly reduced activity of renal vitamin K-dependent gamma-glutamyl carboxylase was observed in the urolithic group as compared with normal controls (P < 0.01).</p><p><b>CONCLUSION</b>It suggests that the reduced carboxylase activity observed in the urolithic patients may play an important role in the course of renal calcium oxalate urolithiasis.</p>

Carbon sources effect on pectinase production from Aspergillus japonicus 586

The effect of different carbon sources on the pectinesterases, endo- and exo-polygalacturonase activities from Aspergillus japonicus 586 was evaluated in liquid media (Manachini solutions) supplemented with different substrate concentrations. The culture medium was inoculated with 5.106 spores/ml and mantained under agitation (140 rpm), at 30ºC, during 122 h. The enzyme evaluation was carried out 24 h after filtration. The crude extract from A. japonicus 586 indicated that the best enzymatic activities were afforded in the presence of 0.5 per cente pectin (pectinesterease), 0.2 per cente pectin and 0.2 per cente glycerol (endopolygalacturonase), and 0.5per cente pectin associated to 0.5 per cente glucose (exopolygalacturonase). Carbon sources concentration, isolated or associated, significantly affects the pectinesterase, and endo- and exopolygalacturonase activities. Pectin, glucose and saccharose, when added to the culture medium in high concentrations, exhibited a repression effect on all the analyzed enzymes.(au)