Under a suitable condition of crystallization, dark brown short rhombohedron crystals could be obtained from FeMoco-deficient MoFe protein (DeltanifE Avl) purified from a nifE deleted mutant DJ35 of Azotobacter vinela...Under a suitable condition of crystallization, dark brown short rhombohedron crystals could be obtained from FeMoco-deficient MoFe protein (DeltanifE Avl) purified from a nifE deleted mutant DJ35 of Azotobacter vinelandii Lipmann grown in NH3-limited medium. The number, size and quality of crystals were significantly affected by either the concentration of precipitants and buffer or diffusion method. The longest sides of the largest crystal of DeltanifE Avl protein, which was obtained by vapor diffusion in the hanging drop method, were 0.12 and 0.13 mm, respectively.展开更多
By incubating the reduced MoFe protein from Azotobacter vinelandii with O phenanthroline under air and chromatographying the incubated solution on Sephadex G 25 column, inactive MoFe protein could be obtained. Its a...By incubating the reduced MoFe protein from Azotobacter vinelandii with O phenanthroline under air and chromatographying the incubated solution on Sephadex G 25 column, inactive MoFe protein could be obtained. Its acetylene reduction activity was remarkably recovered not only by incubation with the reconstituent solution composed of KMnO 4,ferric homocitrate, Na 2S and dithiothreitol,Azotobacter vinelandii with O phenanthroline under air and chromatographying the incubated solution on Sephadex G 25 column, inactive MoFe protein could be obtained. Its acetylene reduction activity was remarkably recovered not only by incubation with the reconstituent solution composed of KMnO 4,ferric homocitrate, Na 2S and dithiothreitol, with O phenanthroline under air and chromatographying the incubated solution on Sephadex G 25 column, inactive MoFe protein could be obtained. Its acetylene reduction activity was remarkably recovered not only by incubation with the reconstituent solution composed of KMnO 4,ferric homocitrate, Na 2S and dithiothreitol, but also with a mixture of 4Fe∶4S clusters and another cluster which had two structure units of 1Mo∶3Fe∶4S bridged by three -OCH 3- at the Mo atoms. Neither the reconstituent solution nor the mixture could reactivate apo MoFe proteins from the mutants deleting nifE and nifH genes and from the mutant UW 45 , which could be reactivated by the FeMoco extracted from the MoFe protein. The results indicated that the FeMoco deficient MoFe proteins from these mutants seemed to be reconstituted only by the clusters which were probably structures only similar to FeMoco. The partially metallocluster deficient MoFe protein could be reconstituted by the clusters with a certain kind of structure and composition; and was changed into different nitrogenase proteins with the ability to fix nitrogen.展开更多
The MoOx/AuNPs composite film modified glassy carbon electrode was fabricated by electro-depositing simultaneously gold nanoparticles and molybdenum oxides using cyclic voltammetry. The morphology and topography of th...The MoOx/AuNPs composite film modified glassy carbon electrode was fabricated by electro-depositing simultaneously gold nanoparticles and molybdenum oxides using cyclic voltammetry. The morphology and topography of the MoOx/AuNPs composite were char-acterized by scan electron microscopy and X-ray photoelectron spectroscopy respectively, and the electrocatalytic oxidation of glucose at the MoOx/AuNPs composite film was inves-tigated and analyzed in detail. It was shown that the MoOx/AuNPs composite was of strong electrocatalytic activity towards oxidation of glucose as well as other saccharides, so that an attempt was made for direct voltammetric determination of glucose. Then the positive scan polarization reverse catalytic voltammetry was proposed for the first time. Based on this method, the pure oxidation current was extracted by subtraction of the blank current in the reverse scan. The current sensitivity was enhanced tremendously and the signal to noise ra-tio was improved adequately. The electrocatalytic oxidation of glucose at the MoOx/AuNPs modified electrode was performed in alkaline medium, a wide linear range from 0.01 mmol/L to 4.0 mmol/L of glucose, a higher current sensitivity of 2.35 mA/（mmol/L·cm2）, and a lower limit of detection of 9.01 μmol/L （at signal/noise=3） were achieved. In addition, the electrocatalytic oxidation of other saccharides such as lactose, fructose and sucrose was also evaluated.展开更多
【目的】微小杆菌属(Exiguobacterium)细菌广泛分布于海洋及非海洋环境中,具有多种代谢途径以适应复杂多样的生境。本研究从能量代谢途径角度出发,探究该属菌株对不同生境的适应能力。【方法】从美国国家生物科技数据中心(National Cent...【目的】微小杆菌属(Exiguobacterium)细菌广泛分布于海洋及非海洋环境中,具有多种代谢途径以适应复杂多样的生境。本研究从能量代谢途径角度出发,探究该属菌株对不同生境的适应能力。【方法】从美国国家生物科技数据中心(National Center for Biotechnology Information, NCBI)数据库中获取146个Exiguobacterium属菌株的基因组,查找并统计光营养、厌氧呼吸和底物代谢等多种能量代谢途径的关键蛋白或关键酶基因在各菌株基因组中的分布,包括光营养型的视紫红质基因、厌氧呼吸营养型的钼辅因子合成蛋白基因,以及底物代谢营养型中乙醛酸分流途径的异柠檬酸裂解酶及苹果酸合酶基因等。根据对应的氨基酸序列构建视紫红质、MoaC和异柠檬酸裂解酶的系统发育树,分析不同能量代谢途径在该属菌株进化过程中的保守性,推测其对于该属菌株的重要性。【结果】Exiguobacterium属中50%的种具有视紫红质基因,其中分离自非海洋生境的菌株更趋向于含有视紫红质基因。本研究所统计的全部非海洋生境菌株中,含有视紫红质基因的菌株占比约为70%,而在海洋生境菌株中该比例仅为19%。Exiguobacterium属约27%的种存在钼辅因子合成蛋白基因,分离自海洋环境的菌株有该类蛋白的可能性更高(32%:21%),具有完整钼辅因子合成途径的菌株集中于进化树同一分支上的少数种。该属约61%的种存在乙醛酸分流途径相关酶基因,这些种隶属进化树的同一分支,且种内所有菌株都具备相关基因,表明乙醛酸分流途径在Exiguobacterium属的分布具有种特异性。【结论】Exiguobacterium属细菌具有多种能量代谢途径相关基因,包括基于视紫红质的光营养型能量代谢途径、基于钼酶的厌氧呼吸型能量代谢途径和底物代谢营养型能量代谢途径中的乙醛酸分流途径。能量代谢途径多样性可能是Exiguobacterium属细菌适应复杂多样生境的机制之一。此外,本研究发现Exiguobacterium属中不同种、同种不同菌株间存在能量代谢途径差异,且能量代谢途径在该属内的分布多不具备种特异性,表明仅通过16S rRNA基因种属鉴定来预测目标菌株的代谢类型可能有较大偏差和局限性。展开更多
文摘Under a suitable condition of crystallization, dark brown short rhombohedron crystals could be obtained from FeMoco-deficient MoFe protein (DeltanifE Avl) purified from a nifE deleted mutant DJ35 of Azotobacter vinelandii Lipmann grown in NH3-limited medium. The number, size and quality of crystals were significantly affected by either the concentration of precipitants and buffer or diffusion method. The longest sides of the largest crystal of DeltanifE Avl protein, which was obtained by vapor diffusion in the hanging drop method, were 0.12 and 0.13 mm, respectively.
文摘By incubating the reduced MoFe protein from Azotobacter vinelandii with O phenanthroline under air and chromatographying the incubated solution on Sephadex G 25 column, inactive MoFe protein could be obtained. Its acetylene reduction activity was remarkably recovered not only by incubation with the reconstituent solution composed of KMnO 4,ferric homocitrate, Na 2S and dithiothreitol,Azotobacter vinelandii with O phenanthroline under air and chromatographying the incubated solution on Sephadex G 25 column, inactive MoFe protein could be obtained. Its acetylene reduction activity was remarkably recovered not only by incubation with the reconstituent solution composed of KMnO 4,ferric homocitrate, Na 2S and dithiothreitol, with O phenanthroline under air and chromatographying the incubated solution on Sephadex G 25 column, inactive MoFe protein could be obtained. Its acetylene reduction activity was remarkably recovered not only by incubation with the reconstituent solution composed of KMnO 4,ferric homocitrate, Na 2S and dithiothreitol, but also with a mixture of 4Fe∶4S clusters and another cluster which had two structure units of 1Mo∶3Fe∶4S bridged by three -OCH 3- at the Mo atoms. Neither the reconstituent solution nor the mixture could reactivate apo MoFe proteins from the mutants deleting nifE and nifH genes and from the mutant UW 45 , which could be reactivated by the FeMoco extracted from the MoFe protein. The results indicated that the FeMoco deficient MoFe proteins from these mutants seemed to be reconstituted only by the clusters which were probably structures only similar to FeMoco. The partially metallocluster deficient MoFe protein could be reconstituted by the clusters with a certain kind of structure and composition; and was changed into different nitrogenase proteins with the ability to fix nitrogen.
文摘The MoOx/AuNPs composite film modified glassy carbon electrode was fabricated by electro-depositing simultaneously gold nanoparticles and molybdenum oxides using cyclic voltammetry. The morphology and topography of the MoOx/AuNPs composite were char-acterized by scan electron microscopy and X-ray photoelectron spectroscopy respectively, and the electrocatalytic oxidation of glucose at the MoOx/AuNPs composite film was inves-tigated and analyzed in detail. It was shown that the MoOx/AuNPs composite was of strong electrocatalytic activity towards oxidation of glucose as well as other saccharides, so that an attempt was made for direct voltammetric determination of glucose. Then the positive scan polarization reverse catalytic voltammetry was proposed for the first time. Based on this method, the pure oxidation current was extracted by subtraction of the blank current in the reverse scan. The current sensitivity was enhanced tremendously and the signal to noise ra-tio was improved adequately. The electrocatalytic oxidation of glucose at the MoOx/AuNPs modified electrode was performed in alkaline medium, a wide linear range from 0.01 mmol/L to 4.0 mmol/L of glucose, a higher current sensitivity of 2.35 mA/（mmol/L·cm2）, and a lower limit of detection of 9.01 μmol/L （at signal/noise=3） were achieved. In addition, the electrocatalytic oxidation of other saccharides such as lactose, fructose and sucrose was also evaluated.
文摘【目的】微小杆菌属(Exiguobacterium)细菌广泛分布于海洋及非海洋环境中,具有多种代谢途径以适应复杂多样的生境。本研究从能量代谢途径角度出发,探究该属菌株对不同生境的适应能力。【方法】从美国国家生物科技数据中心(National Center for Biotechnology Information, NCBI)数据库中获取146个Exiguobacterium属菌株的基因组,查找并统计光营养、厌氧呼吸和底物代谢等多种能量代谢途径的关键蛋白或关键酶基因在各菌株基因组中的分布,包括光营养型的视紫红质基因、厌氧呼吸营养型的钼辅因子合成蛋白基因,以及底物代谢营养型中乙醛酸分流途径的异柠檬酸裂解酶及苹果酸合酶基因等。根据对应的氨基酸序列构建视紫红质、MoaC和异柠檬酸裂解酶的系统发育树,分析不同能量代谢途径在该属菌株进化过程中的保守性,推测其对于该属菌株的重要性。【结果】Exiguobacterium属中50%的种具有视紫红质基因,其中分离自非海洋生境的菌株更趋向于含有视紫红质基因。本研究所统计的全部非海洋生境菌株中,含有视紫红质基因的菌株占比约为70%,而在海洋生境菌株中该比例仅为19%。Exiguobacterium属约27%的种存在钼辅因子合成蛋白基因,分离自海洋环境的菌株有该类蛋白的可能性更高(32%:21%),具有完整钼辅因子合成途径的菌株集中于进化树同一分支上的少数种。该属约61%的种存在乙醛酸分流途径相关酶基因,这些种隶属进化树的同一分支,且种内所有菌株都具备相关基因,表明乙醛酸分流途径在Exiguobacterium属的分布具有种特异性。【结论】Exiguobacterium属细菌具有多种能量代谢途径相关基因,包括基于视紫红质的光营养型能量代谢途径、基于钼酶的厌氧呼吸型能量代谢途径和底物代谢营养型能量代谢途径中的乙醛酸分流途径。能量代谢途径多样性可能是Exiguobacterium属细菌适应复杂多样生境的机制之一。此外,本研究发现Exiguobacterium属中不同种、同种不同菌株间存在能量代谢途径差异,且能量代谢途径在该属内的分布多不具备种特异性,表明仅通过16S rRNA基因种属鉴定来预测目标菌株的代谢类型可能有较大偏差和局限性。
