A mulfistep conversion system composed of phenol hydroxylase （PHrND） and 2,3-dihydroxy-biphenyl 1,2-dioxygenase （BphCLA_4） was used to synthesize methylcatechols and semialdehydes from o- and m-cresol for the firs...A mulfistep conversion system composed of phenol hydroxylase （PHrND） and 2,3-dihydroxy-biphenyl 1,2-dioxygenase （BphCLA_4） was used to synthesize methylcatechols and semialdehydes from o- and m-cresol for the first time. Docking studies displayed by PyMOL predicted that cresols and methylcatechols could be theoretically transformed by this multistep conversion system~ High performance liquid chromatography mass spectrometry （HPLC-MS） analysis also indicated that the products formed from multistep conversion were the corresponding 3-methylcatechol, 4-methylcatechol, 2- hydroxy-3-methyl-6-oxohexa-2,4-dienoic acid （2- hydroxy-3-methyl-ODA） and 2-hydroxy-5-methyl-6-oxo- hexa-2,4-dienoic acid （2-hydroxy-5-methyl-ODA）. The optimal cell concentrations of the recombinant E. coli strain BL21 （DE3） expressing phenol hydroxylase （PHrND） and 2,3-dihydroxy-biphenyl 1,2-dioxygenase （BphCLA_4） and pH for the multistep conversion of o- and m-cresol were 4.0 （g-L-1 cell dry weight） and pH 8.0, respectively. For the first step conversion, the formation rate of 3- methylcatechol （0.29μmol·L-1·min-1·mg-1cell dry weight） from o-cresol was similarly with that ofmethylca- techols （0.28 μmol·L-1·min-1·mg-1 cell dry weight） from m-cresol by strain PHrND. For the second step conversion, strain BphCLA_4 showed higher formation rate （0.83 μmol·L-1·min-1·mg-1 cell dry weight） for 2-hydroxy-3-methyl- ODA and 2-hydroxy-5-methyl-ODA from m-cresol, which was 1.1-fold higher than that for 2-hydroxy-3-methyl- ODA （0.77 μmol·L-1·min-1·mg-1. mglcell dry weight） from ocresol. The present study suggested the potential application of the multistep conversion system for the production of chemical synthons and high-value products.展开更多
Sphingomonas xenophaga QYY, capable of growing significantly on more than ten kinds of aromatic compounds as sole carbon source, was used to study characterization of catechol 1,2-dioxygenase (C12O) in cell extracts. ...Sphingomonas xenophaga QYY, capable of growing significantly on more than ten kinds of aromatic compounds as sole carbon source, was used to study characterization of catechol 1,2-dioxygenase (C12O) in cell extracts. Characterization of the crude C12O showed that the maximum activity was obtained at 40–70°C and pH 7.8–8.8. Metal ions had different influences on the activity of crude C12O. It was suggested that strain QYY possessed an inducible and ferric-dependent C12O. Kinetic studies showed that the value of V max and K m was 0.25 μmol catechol/L/mg protein/min and 52.85 μmol/L, respectively. In addition, the partial purification of C12O was achieved by a HiTrap Q Sepharose column chromatography.展开更多
芝麻是八大类食物过敏原之一,快速准确识别芝麻过敏原对预防其过敏有重要意义。核酸适配体可以高效识别靶标过敏原,在过敏原检测中有良好的应用前景。为了获得芝麻主要过敏原Ses i 2的特异性核酸适体,本研究以Ses i 2为靶标,通过磁珠筛...芝麻是八大类食物过敏原之一,快速准确识别芝麻过敏原对预防其过敏有重要意义。核酸适配体可以高效识别靶标过敏原,在过敏原检测中有良好的应用前景。为了获得芝麻主要过敏原Ses i 2的特异性核酸适体,本研究以Ses i 2为靶标,通过磁珠筛选法(磁珠-SELEX)开展10轮筛选,经由高通量测序获得6条候补序列(S1~S6),并进行家族性、同源性分析及二级结构预测。结果表明,6条候选核酸适体的重复率可达46.38%,其自由能在-9.02到-2.47 kcal·moL^(-1)之间,根据自由能能量稳定原则,S1和S5吉布斯自由能最低最稳定,分别为-6.70和-9.02 kcal·moL^(-1)。利用ELISA试验进行亲和力测试,结果表明核酸适体S1和S2的亲和能力较强,S1:KD=67.02 nmol·L^(-1),R2=0.925 8,S2:KD=97.65 nmol·L^(-1),R2=0.795 1。核酸适体S1与过敏原Ses i 2的结合力和其他过敏原蛋白相比有显著差异,可视为具有特异性。本研究最终获得一条兼具良好亲和力和特异性的核酸适体S1,为芝麻过敏原快速检测提供了技术支撑。展开更多
目的了解中老年2型糖尿病(type 2 diabetes mellitus,T2DM)患者少肌性肥胖(sarcopenic obesity,SO)患病率及不同诊断方法之间的一致性。方法采用系统随机抽样法选取2016年1月至2018年3月于北京地区9家医院内分泌科就诊的≥50岁T2DM患者...目的了解中老年2型糖尿病(type 2 diabetes mellitus,T2DM)患者少肌性肥胖(sarcopenic obesity,SO)患病率及不同诊断方法之间的一致性。方法采用系统随机抽样法选取2016年1月至2018年3月于北京地区9家医院内分泌科就诊的≥50岁T2DM患者,使用生物阻抗法进行身体成分检测;根据2022年欧洲临床营养与代谢学会(European Society for Clinical Nutrition and Metabolism,ESPEN)和欧洲肥胖研究协会(European Association for the Study of Obesity,EASO)方法定义SO,另外3种方法通过肌少症和肥胖的组合进行诊断。肌少症使用2019年亚洲肌少症工作组(Asian Working Group for Sarcopenia,AWGS)建立的标准来定义,肥胖通过体脂(percent of body fat,PBF)、腰围(waist circumference,WC)和体质量指数(body mass index,BMI)来定义。卡方检验进行率的比较,Cohens kappa统计分析比较4种方法的诊断一致性。结果共纳入1125例T2DM受试者,男性586例,年龄[61.2(55.3,67.4)]岁;女性539例,年龄[62.0(56.3,68.1)岁]。使用ESPEN/EASO共识、AWGS+PBF、AWGS+WC和AWGS+BMI标准,中老年T2DM患者SO患病率分别为41.6%、20.4%、30.1%和18.8%。4种方法之间的诊断一致性存在异质性(κ:0.109~0.655)。ESPEN/EASO共识与AWGS+PBF诊断一致性良好(κ:0.655),AWGS+体脂与AWGS+BMI诊断一致性良好(κ:0.637),AWGS+WC与AWGS+BMI(κ:0.359)、与AWGS+PBF诊断一致性中等(κ:0.330)。结论中老年T2DM患者SO患病率高,患病率和诊断一致性在不同诊断方法中存在差异,ESPEN/EASO的共识诊断率最高,AWGS+BMI诊断率最低,ESPEN/EASO共识与AWGS+体脂具有良好的诊断一致性。展开更多
基金The authors gratefully acknowledge the financial supports from the National Natural Science Foundation of China （Grant Nos. 51078054, 51108120, and 51178139） and the National Creative Research Group from the National Natural Science Foundation of China （No. 51121062）.
文摘A mulfistep conversion system composed of phenol hydroxylase （PHrND） and 2,3-dihydroxy-biphenyl 1,2-dioxygenase （BphCLA_4） was used to synthesize methylcatechols and semialdehydes from o- and m-cresol for the first time. Docking studies displayed by PyMOL predicted that cresols and methylcatechols could be theoretically transformed by this multistep conversion system~ High performance liquid chromatography mass spectrometry （HPLC-MS） analysis also indicated that the products formed from multistep conversion were the corresponding 3-methylcatechol, 4-methylcatechol, 2- hydroxy-3-methyl-6-oxohexa-2,4-dienoic acid （2- hydroxy-3-methyl-ODA） and 2-hydroxy-5-methyl-6-oxo- hexa-2,4-dienoic acid （2-hydroxy-5-methyl-ODA）. The optimal cell concentrations of the recombinant E. coli strain BL21 （DE3） expressing phenol hydroxylase （PHrND） and 2,3-dihydroxy-biphenyl 1,2-dioxygenase （BphCLA_4） and pH for the multistep conversion of o- and m-cresol were 4.0 （g-L-1 cell dry weight） and pH 8.0, respectively. For the first step conversion, the formation rate of 3- methylcatechol （0.29μmol·L-1·min-1·mg-1cell dry weight） from o-cresol was similarly with that ofmethylca- techols （0.28 μmol·L-1·min-1·mg-1 cell dry weight） from m-cresol by strain PHrND. For the second step conversion, strain BphCLA_4 showed higher formation rate （0.83 μmol·L-1·min-1·mg-1 cell dry weight） for 2-hydroxy-3-methyl- ODA and 2-hydroxy-5-methyl-ODA from m-cresol, which was 1.1-fold higher than that for 2-hydroxy-3-methyl- ODA （0.77 μmol·L-1·min-1·mg-1. mglcell dry weight） from ocresol. The present study suggested the potential application of the multistep conversion system for the production of chemical synthons and high-value products.
基金Supported by the National Natural Science Foundation of China (Grant No. 50608011)the 39th Postdoctoral Funds of China (Grant No. 20060390983)
文摘Sphingomonas xenophaga QYY, capable of growing significantly on more than ten kinds of aromatic compounds as sole carbon source, was used to study characterization of catechol 1,2-dioxygenase (C12O) in cell extracts. Characterization of the crude C12O showed that the maximum activity was obtained at 40–70°C and pH 7.8–8.8. Metal ions had different influences on the activity of crude C12O. It was suggested that strain QYY possessed an inducible and ferric-dependent C12O. Kinetic studies showed that the value of V max and K m was 0.25 μmol catechol/L/mg protein/min and 52.85 μmol/L, respectively. In addition, the partial purification of C12O was achieved by a HiTrap Q Sepharose column chromatography.
文摘芝麻是八大类食物过敏原之一,快速准确识别芝麻过敏原对预防其过敏有重要意义。核酸适配体可以高效识别靶标过敏原,在过敏原检测中有良好的应用前景。为了获得芝麻主要过敏原Ses i 2的特异性核酸适体,本研究以Ses i 2为靶标,通过磁珠筛选法(磁珠-SELEX)开展10轮筛选,经由高通量测序获得6条候补序列(S1~S6),并进行家族性、同源性分析及二级结构预测。结果表明,6条候选核酸适体的重复率可达46.38%,其自由能在-9.02到-2.47 kcal·moL^(-1)之间,根据自由能能量稳定原则,S1和S5吉布斯自由能最低最稳定,分别为-6.70和-9.02 kcal·moL^(-1)。利用ELISA试验进行亲和力测试,结果表明核酸适体S1和S2的亲和能力较强,S1:KD=67.02 nmol·L^(-1),R2=0.925 8,S2:KD=97.65 nmol·L^(-1),R2=0.795 1。核酸适体S1与过敏原Ses i 2的结合力和其他过敏原蛋白相比有显著差异,可视为具有特异性。本研究最终获得一条兼具良好亲和力和特异性的核酸适体S1,为芝麻过敏原快速检测提供了技术支撑。
文摘目的了解中老年2型糖尿病(type 2 diabetes mellitus,T2DM)患者少肌性肥胖(sarcopenic obesity,SO)患病率及不同诊断方法之间的一致性。方法采用系统随机抽样法选取2016年1月至2018年3月于北京地区9家医院内分泌科就诊的≥50岁T2DM患者,使用生物阻抗法进行身体成分检测;根据2022年欧洲临床营养与代谢学会(European Society for Clinical Nutrition and Metabolism,ESPEN)和欧洲肥胖研究协会(European Association for the Study of Obesity,EASO)方法定义SO,另外3种方法通过肌少症和肥胖的组合进行诊断。肌少症使用2019年亚洲肌少症工作组(Asian Working Group for Sarcopenia,AWGS)建立的标准来定义,肥胖通过体脂(percent of body fat,PBF)、腰围(waist circumference,WC)和体质量指数(body mass index,BMI)来定义。卡方检验进行率的比较,Cohens kappa统计分析比较4种方法的诊断一致性。结果共纳入1125例T2DM受试者,男性586例,年龄[61.2(55.3,67.4)]岁;女性539例,年龄[62.0(56.3,68.1)岁]。使用ESPEN/EASO共识、AWGS+PBF、AWGS+WC和AWGS+BMI标准,中老年T2DM患者SO患病率分别为41.6%、20.4%、30.1%和18.8%。4种方法之间的诊断一致性存在异质性(κ:0.109~0.655)。ESPEN/EASO共识与AWGS+PBF诊断一致性良好(κ:0.655),AWGS+体脂与AWGS+BMI诊断一致性良好(κ:0.637),AWGS+WC与AWGS+BMI(κ:0.359)、与AWGS+PBF诊断一致性中等(κ:0.330)。结论中老年T2DM患者SO患病率高,患病率和诊断一致性在不同诊断方法中存在差异,ESPEN/EASO的共识诊断率最高,AWGS+BMI诊断率最低,ESPEN/EASO共识与AWGS+体脂具有良好的诊断一致性。
