Characterization of the gene for the Fe-protein of the vanadium dependent alternative nitrogenase of Azotobacter vinelandii and construction of a Tn5 mutant |
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Authors: | Ramesh Raina M Amarender Reddy Debabrota Ghosal and H K Das |
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Institution: | (1) Genetic Engineering Unit, School of Environmental Sciences, Jawaharlal Nehru University, 110067 New Delhi, India |
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Abstract: | Summary A sequence homologous to the conventional nifH gene has been cloned from a different region of the Azotobacter vinelandii genome. Tn5 insertions were obtained in this clone and the mutagenized plasmid was used for marker exchange with A. vinelandii strain CA12 ( nifHDK) to obtain Tn5 mutants. These mutants exhibited a Nif- phenotype in the presence of vanadium, unlike CA12 which was Nif+ on vanadium-containing medium. The gene in the cloned nifH-like region is therefore apparently involved in the vanadium dependent alternative pathway of nitrogen fixation. This gene, nifH2, has been sequenced and encodes a protein of 289 amino acids that is similar to nifH in nucleotide sequence, deduced amino acid sequence, predicted secondary structure and hydrophobicity profile. A second open reading frame downstream of nifH2 codes for a protein of 64 amino acids, similar to the ferredoxin (Fd)-like protein encoded downstream of nifH
* in A. chroococcum. Sequence analysis suggests that the nifH2 and Fd-like genes are in a single operon. |
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Keywords: | Azotobacter vinelandii Vanadium nitrogenase nifH2 Marker exchange DNA sequencing |
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