Role of GlnK in NifL-Mediated Regulation of NifA Activity in<i>Azotobacter vinelandii</i>

Rudnick, Paul; Kunz, Christopher; Gunatilaka, Malkanthi K.; Hines, Eric R.; Kennedy, Christina

doi:10.1128/jb.184.3.812-820.2002

Cited by 57 publications

(64 citation statements)

References 47 publications

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“…This study demonstrates that uridylylation of Av GlnK inhibits the interaction with NifL, and we have shown previously that uridylylation of a single subunit within the GlnK trimer is sufficient to decrease the inhibitory influence of NifL on NifA activity (9). In vivo studies also suggest that the uridylylated form of Av GlnK is required to prevent NifL from inactivating NifA under conditions of nitrogen limitation, and interaction between Av GlnK and NifL has been demonstrated recently using a yeast two-hybrid system (41).…”

Section: Discussionmentioning

confidence: 77%

Direct Interaction of the NifL Regulatory Protein with the GlnK Signal Transducer Enables the Azotobacter vinelandiiNifL-NifA Regulatory System to Respond to Conditions Replete for Nitrogen

Little

Colombo

Leech

et al. 2002

Journal of Biological Chemistry

107

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Section: Discussionmentioning

confidence: 77%

Direct Interaction of the NifL Regulatory Protein with the GlnK Signal Transducer Enables the Azotobacter vinelandiiNifL-NifA Regulatory System to Respond to Conditions Replete for Nitrogen

Little

Colombo

Leech

et al. 2002

Journal of Biological Chemistry

107

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“…The GlnK-NifL-NifA ternary complex is formed under nitrogen-excess conditions when GlnK is primarily in the nonuridylylated form. Uridylylation of GlnK under nitrogen-limiting conditions prevents this interaction (17)(18)(19). We infer that the R306C substitution locks NifL in a conformation that is analogous to that shown in C and D, so that it is competent to inhibit NifA irrespective of other signals.…”

Section: Discussionmentioning

confidence: 87%

“…Unlike the HPKs, the GHKL domain of NifL does not exhibit ATP hydrolysis or transphosphorylation activity, but the binding of ADP to this domain strongly stimulates the inhibitory activity of NifL and the stability of the NifL-NifA complex (8,15,16). Also, the GHKL domain is involved in sensing the nitrogen status because it is the site of interaction with the signal transduction protein GlnK (17)(18)(19), the PII-like protein of A. vinelandii (20)(21)(22)(23).…”

mentioning

confidence: 99%

A crucial arginine residue is required for a conformational switch in NifL to regulate nitrogen fixation in Azotobacter vinelandii

Martínez‐Argudo

Little

Dixon

2004

Proc. Natl. Acad. Sci. U.S.A.

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NifL is an antiactivator that tightly regulates transcription of genes required for nitrogen fixation in Azotobacter vinelandii by controlling the activity of its partner protein NifA, a member of the family of 54 -dependent transcriptional activators. Although the C-terminal region of A. vinelandii NifL shows homology to the transmitter domains of histidine protein kinases, signal transduction between NifL and NifA is conveyed by means of proteinprotein interactions rather than by phosphorylation. Binding of the ligand 2-oxoglutarate to NifA plays a crucial role in preventing inhibition by NifL under conditions appropriate for nitrogen fixation. We have used a suppressor screen to identify a critical arginine residue (R306) in NifL that is required to release NifA from inhibition under appropriate environmental conditions. Amino acid substitutions at position 306 result in constitutive inhibition of NifA activity by NifL, thus preventing nitrogen fixation. Biochemical studies with one of the mutant proteins demonstrate that the substitution alters the conformation of NifL significantly and prevents the response of NifA to 2-oxoglutarate. We propose that arginine 306 is critical for the propagation of signals perceived by A. vinelandii NifL in response to the redox and fixed-nitrogen status and is required for a conformational switch that inactivates the inhibitory function of NifL under conditions appropriate for nitrogen fixation.2-oxoglutarate ͉ signal transduction ͉ antiactivator ͉ redox control ͉ nitrogen regulation S tructural rearrangements of sensor proteins in response to environmental cues provide a fundamental mechanism for signal propagation within cells. However, although conformational changes have been well characterized in isolated signaling domains, mechanisms for signal transmission by means of interdomain interactions are frequently less well understood. For example, structural studies have identified ligand-induced conformational changes in the sensor domains of histidine protein kinases (HPKs), but it is not known how these changes are communicated to the kinase domain to control phosphoryl transfer.The Azotobacter vinelandii NifL regulatory protein is a histidine kinase-like protein that controls the expression of the genes required for nitrogen fixation in response to the redox, nitrogen, and carbon status. NifL is an antiactivator that tightly regulates the activity of its partner protein NifA, a member of the family of 54 -transcriptional activators (1, 2), by means of the formation of an inhibitory complex (3-5). The domain architecture of NifL is similar to that of some HPKs, with an N-terminal Per-ArntSim (PAS) domain (6, 7) containing a flavin adenine dinucleotide (FAD) cofactor that senses the redox status (8, 9) and a C-terminal domain containing conserved residues corresponding to the N, G1, F, and G2 boxes that constitute the ATPbinding domain of the GHKL superfamily of ATPases (10-14) (Fig. 1). Unlike the HPKs, the GHKL domain of NifL does not exhibit ATP hydrolysis or transphos...

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“…It has been successfully used to explore binding between several components of the ntr and nif regulatory systems (Lei et al, 1999;Martínez-Argudo et al, 2002;Rudnick et al, 2002;Pawlowski et al, 2003;Chen et al, 2005). Thus, a direct protein-protein binding was detected between NifA and NifL in K. pneumoniae, Azotobacter vinelandii and Enterobacter cloacae (Lei et al, 1999;Martínez-Argudo et al, 2002;Liao et al, 2002).…”

Section: Discussionmentioning

confidence: 99%

Interaction between NifL and NifA in the nitrogen-fixing Pseudomonas stutzeri A1501

et al. 2006

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Pseudomonas stutzeri strain A1501 isolated from rice fixes nitrogen under microaerobic conditions in the free-living state. This paper describes the properties of nifL and nifA mutants as well as the physical interaction between NifL and NifA proteins. A nifL mutant strain that carried a mutation nonpolar on nifA expression retained nitrogenase activity. Complementation with a plasmid containing only nifL led to a decrease in nitrogenase activity in both the wild-type and the nifL mutant, suggesting that NifL acts as an antiactivator of NifA activity. Using the yeast two-hybrid system and purified protein domains of NifA and NifL, an interaction was shown between the C-terminal domain of NifL and the central domain of NifA, suggesting that NifL antiactivator activity is mediated by direct protein interaction with NifA.

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Role of GlnK in NifL-Mediated Regulation of NifA Activity inAzotobacter vinelandii

Cited by 57 publications

References 47 publications

Direct Interaction of the NifL Regulatory Protein with the GlnK Signal Transducer Enables the Azotobacter vinelandiiNifL-NifA Regulatory System to Respond to Conditions Replete for Nitrogen

Direct Interaction of the NifL Regulatory Protein with the GlnK Signal Transducer Enables the Azotobacter vinelandiiNifL-NifA Regulatory System to Respond to Conditions Replete for Nitrogen

A crucial arginine residue is required for a conformational switch in NifL to regulate nitrogen fixation in Azotobacter vinelandii

Interaction between NifL and NifA in the nitrogen-fixing Pseudomonas stutzeri A1501

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