PMID:23144248

Citation	Moure, VR, Danyal, K, Yang, ZY, Wendroth, S, Müller-Santos, M, Pedrosa, FO, Scarduelli, M, Gerhardt, EC, Huergo, LF, Souza, EM and Seefeldt, LC (2013) The nitrogenase regulatory enzyme dinitrogenase reductase ADP-ribosyltransferase (DraT) is activated by direct interaction with the signal transduction protein GlnB. J. Bacteriol. 195:279-86
Abstract	Fe protein (dinitrogenase reductase) activity is reversibly inactivated by dinitrogenase reductase ADP-ribosyltransferase (DraT) in response to an increase in the ammonium concentration or a decrease in cellular energy in Azospirillum brasilense, Rhodospirillum rubrum, and Rhodobacter capsulatus. The ADP-ribosyl is removed by the dinitrogenase reductase-activating glycohydrolase (DraG), promoting Fe protein reactivation. The signaling pathway leading to DraT activation by ammonium is still not completely understood, but the available evidence shows the involvement of direct interaction between the enzyme and the nitrogen-signaling P(II) proteins. In A. brasilense, two P(II) proteins, GlnB and GlnZ, were identified. We used Fe protein from Azotobacter vinelandii as the substrate to assess the activity of A. brasilense DraT in vitro complexed or not with P(II) proteins. Under our conditions, GlnB was necessary for DraT activity in the presence of Mg-ADP. The P(II) effector 2-oxoglutarate, in the presence of Mg-ATP, inhibited DraT-GlnB activity, possibly by inducing complex dissociation. DraT was also activated by GlnZ and by both uridylylated P(II) proteins, but not by a GlnB variant carrying a partial deletion of the T loop. Kinetics studies revealed that the A. brasilense DraT-GlnB complex was at least 18-fold more efficient than DraT purified from R. rubrum, but with a similar K(m) value for NAD(+). Our results showed that ADP-ribosylation of the Fe protein does not affect the electronic state of its metal cluster and prevents association between the Fe and MoFe proteins, thus inhibiting electron transfer.
Links	PubMed PMC3553828 Online version:10.1128/JB.01517-12
Keywords	ADP Ribose Transferases/isolation & purification; ADP Ribose Transferases/metabolism; Adenosine Diphosphate/metabolism; Azospirillum brasilense/enzymology; Azotobacter vinelandii/enzymology; Bacterial Proteins/metabolism; Coenzymes; Enzyme Inhibitors/metabolism; Ketoglutaric Acids/metabolism; Kinetics; Magnesium/metabolism; NAD/metabolism; Oxidoreductases/isolation & purification; Oxidoreductases/metabolism; PII Nitrogen Regulatory Proteins/metabolism; Protein Binding
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
RHORU:DRAT		GO:0006471: protein ADP-ribosylation	ECO:0000314:		P		Figure 2 shows activation and interaction with different PII proteins that form in vitro complexes and the Fe protein. Fe protein activity, and thus, DraT activity was measured by the evolution of hydrogen.	complete CACAO 9022
AZOBR:GLNB		GO:0006808: regulation of nitrogen utilization	ECO:0000314:		P		Figure 1. Preincubation of the Fe protein with DraT in the presence of increasing concentrations of GlnB resulted in Fe protein inhibition in a GlnB-dependent manner	complete CACAO 9168

References

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