Ammonium inhibition of nitrogenase activity in Herbaspirillum seropedicae.
The effect of oxygen, ammonium ion, and amino acids on nitrogenase activity in the root-associated N2-fixing bacterium Herbaspirillum seropedicae was investigated in comparison with Azospirillum spp. and Rhodospirillum rubrum. H. seropedicae is microaerophilic, and its optimal dissolved oxygen level is from 0.04 to 0.2 kPa for dinitrogen fixation but higher when it is supplied with fixed nitrogen. No nitrogenase activity was detected when the dissolved O2 level corresponded to 4.0 kPa. Ammonium, a product of the nitrogenase reaction, reversibly inhibited nitrogenase activity when added to derepressed cell cultures. However, the inhibition of nitrogenase activity was only partial even with concentrations of ammonium chloride as high as 20 mM. Amides such as glutamine and asparagine partially inhibited nitrogenase activity, but glutamate did not. Nitrogenase in crude extracts prepared from ammonium-inhibited cells showed activity as high as in extracts from N2-fixing cells. The pattern of the dinitrogenase and the dinitrogenase reductase revealed by the immunoblotting technique did not change upon ammonium chloride treatment of cells in vivo. No homologous sequences were detected with the draT-draG probe from Azospirillum lipoferum. There is no clear evidence that ADP-ribosylation of the dinitrogenase reductase is involved in the ammonium inhibition of H. seropedicae. The uncoupler carbonyl cyanide m-chlorophenylhydrazone decreased the intracellular ATP concentration and inhibited the nitrogenase activity of whole cells. The ATP pool was not significantly disturbed when cultures were treated with ammonium in vivo. Possible mechanisms for inhibition by ammonium of whole-cell nitrogenase activity in H. seropedicae are discussed.
Fu
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Department of Biochemistry, College of Agricultural and Life Sciences, University of Wisconsin-Madison 53706.
Burris
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D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000641:Ammonia / Q000494:pharmacology*; D015153:Blotting, Western; D004269:DNA, Bacterial / Q000235:genetics; D017037:Dinitrogenase Reductase; D005288:Ferredoxins / Q000378:metabolism; D006090:Gram-Negative Bacteria / Q000502:physiology*; D009584:Nitrogen / Q000494:pharmacology; D009586:Nitrogen Fixation; D009591:Nitrogenase / Q000037:antagonists & inhibitors* / Q000378:metabolism; D009693:Nucleic Acid Hybridization; D010100:Oxygen / Q000494:pharmacology
D004269:DNA, Bacterial; D017037:Dinitrogenase Reductase; D005288:Ferredoxins; D000246:Adenosine Diphosphate Ribose; D000641:Ammonia; D000255:Adenosine Triphosphate; D009591:Nitrogenase; D009584:Nitrogen; D010100:Oxygen