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
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.
