Regulation of dinitrogenase reductase ADP-ribosyltransferase and dinitrogenase reductase-activating glycohydrolase by a redox-dependent conformational change of nitrogenase Fe protein.
The nitrogenase-regulating enzymes dinitrogenase reductase ADP-ribosyltransferase (DRAT) and dinitrogenase reductase-activating glycohydrolase (DRAG), from Rhodospirillum rubrum, were shown to be sensitive to the redox status of the [Fe(4)S(4)](1+/2+) cluster of nitrogenase Fe protein from R. rubrum or Azotobacter vinelandii. DRAG had <2% activity with oxidized R. rubrum Fe protein relative to activity with reduced Fe protein. The activity of DRAG with oxygen-denatured Fe protein or a low molecular weight substrate, N(alpha)-dansyl-N(omega)-(1,N(6)-etheno-ADP-ribosyl)-arginine methyl ester, was independent of redox potential. The redox midpoint potential of DRAG activation of Fe protein was -430 mV versus standard hydrogen electrode, coinciding with the midpoint potential of the [Fe(4)S(4)] cluster from R. rubrum Fe protein. DRAT was found to have a specificity opposite that of DRAG, exhibiting low (<20%) activity with 87% reduced R. rubrum Fe protein relative to activity with fully oxidized Fe protein. A mutant of R. rubrum in which the rate of oxidation of Fe protein was substantially decreased had a markedly slower rate of ADP-ribosylation in vivo in response to 10 mM NH(4)Cl or darkness stimulus. It is concluded that the redox state of Fe protein plays a significant role in regulation of the activities of DRAT and DRAG in vivo.
Halbleib
C M
CM
Department of Biochemistry, Center for the Study of Nitrogen Fixation, University of Wisconsin, Madison, Wisconsin 53706, USA.
Zhang
Y
Y
Ludden
P W
PW
D036002:ADP Ribose Transferases / Q000378:metabolism*; D004789:Enzyme Activation; D006026:Glycoside Hydrolases / Q000378:metabolism*; D009154:Mutation; D009699:N-Glycosyl Hydrolases; D009586:Nitrogen Fixation; D009591:Nitrogenase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010084:Oxidation-Reduction; D010088:Oxidoreductases; D011487:Protein Conformation; D012247:Rhodospirillum rubrum / Q000201:enzymology*; D013329:Structure-Activity Relationship
D010088:Oxidoreductases; D009591:Nitrogenase; C029130:nitrogenase reductase; D036002:ADP Ribose Transferases; C057637:dinitrogenase reductase ADP-ribosyltransferase; D006026:Glycoside Hydrolases; D009699:N-Glycosyl Hydrolases; C036091:ADP-ribosyl-(dinitrogen reductase) hydrolase