Background:Very long chain acyl-CoA dehydrogenase deficiency(VLCADD)is an inherited metabolic disease caused by deleterious mutations in the ACADVL gene that encodes very long chain acyl-CoA dehydrogenase(VLCAD),and w...Background:Very long chain acyl-CoA dehydrogenase deficiency(VLCADD)is an inherited metabolic disease caused by deleterious mutations in the ACADVL gene that encodes very long chain acyl-CoA dehydrogenase(VLCAD),and which can present as cardiomyopathy in neonates,as hypoketotic hypoglycemia in infancy,and as myopathy in late-onset patients.Although many ACADVL mutations have been described,no prevalent mutations in the ACADVL gene have been associated with VLCADD.Herein,we report the clinical course of the disease and explore the genetic mutation spectrum in seven Chinese patients with VLCADD.Methods:Seven Chinese patients,from newborn to 17 years old,were included in this study.Tandem mass spectrometry was performed to screen for VLCAD defi ciency.All exons and fl anking introns of the ACADVL gene were analyzed using polymerase chain reaction and direct sequencing.Online analysis tools were used to predict the impact of novel mutations.Results:All cases had elevated serum levels of tetradecanoylcarnitine(C14:1)which is the characteristic biomarker for VLCADD.The phenotype of VLCADD is heterogeneous.Two patients were hospitalized for hypoactivity and hypoglycemia shortly after birth.Three patients showed hepatomegaly and hypoglycemia in infancy.The other two adolescent patients showed initial manifestations of exercise intolerance or rhabdomyolysis.Three of the patients died at the age of 6-8 months.Eleven different mutations in the ACADVL gene in the 7 patients were identified,including seven reported mutations(p.S22X,p.W427X,p.A213T,p.G222R,p.R450H,c.296-297delCA,c.1605+1G>T)and four novel mutations(p.S72F,p.Q100X,p.M437T,p.D466Y).The p.R450H and p.D466Y(14.28%,2/14 alleles)mutations were identifi ed in two alleles respectively.Conclusions:The clinical manifestations were heterogeneous and ACADVL gene mutations were heterozygous in the seven VLCADD Chinese patients.R450H may be a relatively common mutation in Asian populations.The genotype and phenotype had a certain correlation in our patients.展开更多
Genes encoding enzymes involved in biosynthesis of very long chain fatty acids were significantly up-regulatedduring early cotton fiber development. Two cDNAs, GhKCR1 and GhKCR2 encoding putative cotton 3-ketoacyl-CoA...Genes encoding enzymes involved in biosynthesis of very long chain fatty acids were significantly up-regulatedduring early cotton fiber development. Two cDNAs, GhKCR1 and GhKCR2 encoding putative cotton 3-ketoacyl-CoAreductases that catalyze the second step in fatty acid elongation, were isolated from developing cotton fibers. GhKCR1and 2 contain open reading frames of 963 bp and 924 bp encoding proteins of 320 and 307 amino acid residues,respectively. Quantatitive RT-PCR analysis showed that both these genes were highly preferentially expressed duringthe cotton fiber elongation period with much lower levels recovered from roots, stems and leaves. GhKCR1 and 2showed 30%-32% identity to Saccharomyces cerevisiae Ybr159p at the deduced amino acid level. These cotton cDNAswere cloned and expressed in yeast haploid ybr159w? mutant that was deficient in 3-ketoacyl-CoA reductase activity.Wild-type growth rate was restored in ybr159w? cells that expressed either GhKCR1 or 2. Further analysis showed thatGhKCR1 and 2 were co-sedimented within the membranous pellet fraction after high-speed centrifugation, similar to theyeast endoplasmic reticulum marker ScKar2p. Both GhKCR(s) showed NADPH-dependent 3-ketoacyl-CoA reductaseactivity in an in vitro assay system using palmitoyl-CoA and malonyl-CoA as substrates. Our results suggest thatGhKCR1 and 2 are functional orthologues of ScYbr159p.展开更多
NADP(H) dependent retinol dehydrogenase/reductase (NRDR) was an important retinoic acid synthase, which was first purified from rabbit liver in 1997. In order to study the function of the NRDR gene,the full length cDN...NADP(H) dependent retinol dehydrogenase/reductase (NRDR) was an important retinoic acid synthase, which was first purified from rabbit liver in 1997. In order to study the function of the NRDR gene,the full length cDNA of bovine NRDR was cloned. According to the conserved sequences of human, mouse and rabbit NRDR cDNA, a pair of primers was designed to amplify a 294 bp DNA fragment of bovine liver NRDR, and then the full length of NRDR cDNA (AF487454) was cloned by using 3′ RACE and 5′ RACE. All the cloned NRDR proteins consist of 260 amino acid residues and showed high identity among them. The tri peptide of human, mouse and rabbit NRDR C end was SRL and that of bovine NRDR C end was SHL, but both were considered to be peroxisomal target signal 1 (PTS1). RT PCR demonstrated that NRDR gene was expressed in liver, heart, lung, kidney, stomach and intestine, and was not found in pancreas, muscle, artery and skin. The full length bovine NRDR cDNA has been successfully cloned and the sequence was analyzed. It provided a reliable foundation to investigate the biological function of this protein.展开更多
基金supported by grants from the National Natural Science Foundation of China(81170811,30973216)Shanghai School Board(12ZZ114)and Shanghai Health Bureau(20134005)+1 种基金supported by grants from the Major Program of Shanghai Committee of Science and Technology(11dz195030)the National Key Technology R&D Program(2012BAI09B04).
文摘Background:Very long chain acyl-CoA dehydrogenase deficiency(VLCADD)is an inherited metabolic disease caused by deleterious mutations in the ACADVL gene that encodes very long chain acyl-CoA dehydrogenase(VLCAD),and which can present as cardiomyopathy in neonates,as hypoketotic hypoglycemia in infancy,and as myopathy in late-onset patients.Although many ACADVL mutations have been described,no prevalent mutations in the ACADVL gene have been associated with VLCADD.Herein,we report the clinical course of the disease and explore the genetic mutation spectrum in seven Chinese patients with VLCADD.Methods:Seven Chinese patients,from newborn to 17 years old,were included in this study.Tandem mass spectrometry was performed to screen for VLCAD defi ciency.All exons and fl anking introns of the ACADVL gene were analyzed using polymerase chain reaction and direct sequencing.Online analysis tools were used to predict the impact of novel mutations.Results:All cases had elevated serum levels of tetradecanoylcarnitine(C14:1)which is the characteristic biomarker for VLCADD.The phenotype of VLCADD is heterogeneous.Two patients were hospitalized for hypoactivity and hypoglycemia shortly after birth.Three patients showed hepatomegaly and hypoglycemia in infancy.The other two adolescent patients showed initial manifestations of exercise intolerance or rhabdomyolysis.Three of the patients died at the age of 6-8 months.Eleven different mutations in the ACADVL gene in the 7 patients were identified,including seven reported mutations(p.S22X,p.W427X,p.A213T,p.G222R,p.R450H,c.296-297delCA,c.1605+1G>T)and four novel mutations(p.S72F,p.Q100X,p.M437T,p.D466Y).The p.R450H and p.D466Y(14.28%,2/14 alleles)mutations were identifi ed in two alleles respectively.Conclusions:The clinical manifestations were heterogeneous and ACADVL gene mutations were heterozygous in the seven VLCADD Chinese patients.R450H may be a relatively common mutation in Asian populations.The genotype and phenotype had a certain correlation in our patients.
基金supported by grants from China Na-tional Basic Research Program (NO. 2004CB117302)National Natural Science Foundation of China (No.30470171)the Sigrid Jusélius Foundation Finland and the Academy of Finland
文摘Genes encoding enzymes involved in biosynthesis of very long chain fatty acids were significantly up-regulatedduring early cotton fiber development. Two cDNAs, GhKCR1 and GhKCR2 encoding putative cotton 3-ketoacyl-CoAreductases that catalyze the second step in fatty acid elongation, were isolated from developing cotton fibers. GhKCR1and 2 contain open reading frames of 963 bp and 924 bp encoding proteins of 320 and 307 amino acid residues,respectively. Quantatitive RT-PCR analysis showed that both these genes were highly preferentially expressed duringthe cotton fiber elongation period with much lower levels recovered from roots, stems and leaves. GhKCR1 and 2showed 30%-32% identity to Saccharomyces cerevisiae Ybr159p at the deduced amino acid level. These cotton cDNAswere cloned and expressed in yeast haploid ybr159w? mutant that was deficient in 3-ketoacyl-CoA reductase activity.Wild-type growth rate was restored in ybr159w? cells that expressed either GhKCR1 or 2. Further analysis showed thatGhKCR1 and 2 were co-sedimented within the membranous pellet fraction after high-speed centrifugation, similar to theyeast endoplasmic reticulum marker ScKar2p. Both GhKCR(s) showed NADPH-dependent 3-ketoacyl-CoA reductaseactivity in an in vitro assay system using palmitoyl-CoA and malonyl-CoA as substrates. Our results suggest thatGhKCR1 and 2 are functional orthologues of ScYbr159p.
文摘NADP(H) dependent retinol dehydrogenase/reductase (NRDR) was an important retinoic acid synthase, which was first purified from rabbit liver in 1997. In order to study the function of the NRDR gene,the full length cDNA of bovine NRDR was cloned. According to the conserved sequences of human, mouse and rabbit NRDR cDNA, a pair of primers was designed to amplify a 294 bp DNA fragment of bovine liver NRDR, and then the full length of NRDR cDNA (AF487454) was cloned by using 3′ RACE and 5′ RACE. All the cloned NRDR proteins consist of 260 amino acid residues and showed high identity among them. The tri peptide of human, mouse and rabbit NRDR C end was SRL and that of bovine NRDR C end was SHL, but both were considered to be peroxisomal target signal 1 (PTS1). RT PCR demonstrated that NRDR gene was expressed in liver, heart, lung, kidney, stomach and intestine, and was not found in pancreas, muscle, artery and skin. The full length bovine NRDR cDNA has been successfully cloned and the sequence was analyzed. It provided a reliable foundation to investigate the biological function of this protein.
文摘S.tenebrarius H6主要产生安普霉素、氨甲酰妥布霉素和氨甲酰卡那霉素B,其中后两者仅在3′位有差异。为克隆相关基因,根据已报道的aprD3、livY、gentY基因设计兼并性PCR引物,并采用SON-PCR(singleoligonucleotide nested PCR)方法和LASP-PCR(linear amplification single primer PCR)方法,扩增获得部分SCD1基因,Blast分析为短链脱氢酶基因。采用基因阻断技术,使S.tenebrariusH6中SCD1基因失活。与野生菌株相比,变株的抗生素组分和含量几乎没有变化,但变株产孢子能力下降,且产孢子时间推迟,推测该基因与孢子的生成调节相关。
