A mutant UW 3, which is unable to fix N 2 in the presence of Mo (Nif -) but can undergo phenotypic reversal to Nif + under Mo_deficient conditions, was able to grow in Cr_containing but Mo_ and NH 3_deficient...A mutant UW 3, which is unable to fix N 2 in the presence of Mo (Nif -) but can undergo phenotypic reversal to Nif + under Mo_deficient conditions, was able to grow in Cr_containing but Mo_ and NH 3_deficient medium. A partly purified nitrogenase component Ⅰ protein obtained from UW 3 grown on the Cr_containing medium was shown to contain Fe and Cr (atom ratio of Fe to Cr and Mo to Cr: 11.60 and 0.41) and to have 70% of the C 2H 2_ and H +_reduction activity of MoFe protein from the wild_type strain of Azotobacter vinelandii Lipmann. The Cr_containing protein was different in subunit composition from that of MnFe protein purified from the mutant strain grown in the presence of Mn, but similar to that of MoFe protein, that is, it was a tetramer composed of two different subunits (α 2β 2). The preliminary results indicated that the Cr_containing protein might be a nitrogenase component Ⅰ protein.展开更多
The Deltanif E MoFe protein (Deltanif E Av1) was obtained by a chromatography on DE52, Sephacryl S-300 and Q-Sepharose columns from the unheated crude extract of nifE-deleted mutant strain (DJ35) of Azotobacter vinela...The Deltanif E MoFe protein (Deltanif E Av1) was obtained by a chromatography on DE52, Sephacryl S-300 and Q-Sepharose columns from the unheated crude extract of nifE-deleted mutant strain (DJ35) of Azotobacter vinelandii Lipmann. The analysis by SDS-PAGE showed that the Delta nif EAv1 was similar to OP MoFe protein (Av1) of A. vinelandii in the kinds and molecular weights of subunits (alpha and beta subunit). When complemented with nitrogenase Fe protein (M), the A nif EAv1 had hardly any proton-reduction activity, but could be significantly activated by FeMoco extracted from OP Av1. After the Delta nif E Av1 was treated with an excess o-phenanthroline (o-phen) and chromatographied on Sephadex G-25 column under atmosphere of Ar, Delta nif E Av1(C) was obtained. In the presence of both Av2 and MgATP regeneration system, the Delta nif EAv1(C), rather than A nif EAv1, was significantly activated in vitro by a reconstituent solution containing Mn which composed of KMnO4, ferric homocitrate, Na2S, Na2S2O4 (DT) and dithiothreitol (DTT). But in the absence of MgATP or Av2, the activation of Delta nifE Av1(C) did not happen. It indicates that activation of Delta nif EAv1 by RS-Mn requires the pretreatment with o-phen and the simultaneous presence of Av2 and MgATP.展开更多
文摘A mutant UW 3, which is unable to fix N 2 in the presence of Mo (Nif -) but can undergo phenotypic reversal to Nif + under Mo_deficient conditions, was able to grow in Cr_containing but Mo_ and NH 3_deficient medium. A partly purified nitrogenase component Ⅰ protein obtained from UW 3 grown on the Cr_containing medium was shown to contain Fe and Cr (atom ratio of Fe to Cr and Mo to Cr: 11.60 and 0.41) and to have 70% of the C 2H 2_ and H +_reduction activity of MoFe protein from the wild_type strain of Azotobacter vinelandii Lipmann. The Cr_containing protein was different in subunit composition from that of MnFe protein purified from the mutant strain grown in the presence of Mn, but similar to that of MoFe protein, that is, it was a tetramer composed of two different subunits (α 2β 2). The preliminary results indicated that the Cr_containing protein might be a nitrogenase component Ⅰ protein.
文摘The Deltanif E MoFe protein (Deltanif E Av1) was obtained by a chromatography on DE52, Sephacryl S-300 and Q-Sepharose columns from the unheated crude extract of nifE-deleted mutant strain (DJ35) of Azotobacter vinelandii Lipmann. The analysis by SDS-PAGE showed that the Delta nif EAv1 was similar to OP MoFe protein (Av1) of A. vinelandii in the kinds and molecular weights of subunits (alpha and beta subunit). When complemented with nitrogenase Fe protein (M), the A nif EAv1 had hardly any proton-reduction activity, but could be significantly activated by FeMoco extracted from OP Av1. After the Delta nif E Av1 was treated with an excess o-phenanthroline (o-phen) and chromatographied on Sephadex G-25 column under atmosphere of Ar, Delta nif E Av1(C) was obtained. In the presence of both Av2 and MgATP regeneration system, the Delta nif EAv1(C), rather than A nif EAv1, was significantly activated in vitro by a reconstituent solution containing Mn which composed of KMnO4, ferric homocitrate, Na2S, Na2S2O4 (DT) and dithiothreitol (DTT). But in the absence of MgATP or Av2, the activation of Delta nifE Av1(C) did not happen. It indicates that activation of Delta nif EAv1 by RS-Mn requires the pretreatment with o-phen and the simultaneous presence of Av2 and MgATP.
