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Applied and Environmental Microbiology, September 2005, p. 5637-5641, Vol. 71, No. 9
0099-2240/05/$08.00+0     doi:10.1128/AEM.71.9.5637-5641.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

SHORT REPORT

Nitrogenase Switch-Off by Ammonium Ions in Azospirillum brasilense Requires the GlnB Nitrogen Signal-Transducing Protein

Giseli Klassen,2 Emanuel M. Souza,1* M. Geoffrey Yates,1 Liu Un Rigo,1 Roberta M. Costa,2 Juliana Inaba,1 and Fábio O. Pedrosa1*

Departamento de Bioquímica e Biologia Molecular,1 Departamento de Patologia Básica, Universidade Federal do Paraná, Curitiba, Brasil2

Received 3 November 2004/ Accepted 21 March 2005


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ABSTRACT
 
Nitrogenase activity in several diazotrophs is switched off by ammonium and reactivated after consumption. The signaling pathway to this system in Azospirillum brasilense is not understood. We show that ammonium-dependent switch-off through ADP-ribosylation of Fe protein was partial in a glnB mutant of A. brasilense but absent in a glnB glnZ double mutant. Triggering of inactivation by anaerobic conditions was not affected in either mutant. The results suggest that glnB is necessary for full ammonium-dependent nitrogenase switch-off in A. brasilense.


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INTRODUCTION
 
Nitrogenase activity was first shown to be reversibly inhibited by low levels of ammonium ions or energy depletion in Rhodopseudomonas palustris by Zumft and Castillo (40); this effect, called nitrogenase switch-off/switch-on, was later shown for Rhodospirillum rubrum by Ludden and coworkers to be due to the ADP-ribosylation of an arginine residue of one of the Fe protein (dinitrogenase reductase) subunits by the enzyme dinitrogenase reductase ADP-ribosyl transferase (DRAT). Removal of the ADP-ribosyl moiety and reactivation of the Fe protein is catalyzed by the dinitrogenase reductase-activating glycohydrolase (DRAG) (12, 13, 19). Control of nitrogenase activity by ADP-ribosylation in response to the increase of ammonium concentration or energy depletion also occurs in other diazotrophs, such as Rhodobacter capsulatus, Azospirillum brasilense, and Azospirillum lipoferum, most of these belonging to the subclass of alpha-Proteobacteria (9, 10, 12, 25, 39). The exceptions are Azotobacter chroococcum (of gamma-Proteobacteria) and Azoarcus sp. strain BH72 (of beta-Proteobacteria) (20, 21), although the presence of DRAT and DRAG has not been clearly established for these last two bacteria.

The activities of the enzymes DRAG and DRAT in A. brasilense are opposingly regulated depending on the ammonium concentration (35, 36). On addition of ammonium ions to a culture of A. brasilense fixing nitrogen, the DRAT protein is temporarily activated and DRAG is inactivated, resulting in a rapid increase in the ADP-ribosylation of the Fe protein of nitrogenase (35, 36). After ammonium is consumed, the transferase is inactivated and the glycohydrolase restores nitrogenase activity (38, 39). The signal pathway leading to ammonium-induced inhibition or activation of these proteins is dependent on members of the PII family of signal transduction proteins (1, 2). In A. brasilense, signaling of ammonium levels to the DRAT/DRAG system involves a PII paralog, GlnZ, which is necessary for reactivation of the Fe protein following depletion of ammonium levels in ammonium-shocked cultures (15). In R. capsulatus, a glnB mutant fixed nitrogen but inhibition of nitrogenase by ammonium ions was impaired (16) and ADP-ribosylation of the Fe protein in this organism was found to be dependent on the ammonium transporter AmtB (31). In R. rubrum, either GlnB or its paralog GlnJ was required for the regulation of the ADP-ribosylation of the Fe protein (33). Nitrogenase switch-off in Azoarcus sp. strain BH72 was apparently dependent on the AmtB and GlnK proteins but not on GlnB or GlnY (20). In addition, the NtrC protein may be involved in this process in some organisms: NtrC mutants of both A. brasilense and R. rubrum were deficient in ammonium-induced switch-off (23, 32, 37).

Since in many of these organisms the PII protein (GlnB) also participates in the signaling of the N status to the nif gene transcriptional regulatory system (1), this family of proteins seems to play a central role in regulation of nitrogen fixation. Because the GlnB protein is required for A. brasilense NifA (nif gene activator) activity, probably through interaction with the N-terminal domain, glnB mutants of A. brasilense are Nif negative (6, 17, 18). To study the effect of GlnB on nitrogenase switch-off, the above complication was overcome by introducing a plasmid constitutively expressing the N-truncated NifA protein of Herbaspirillum seropedicae in A. brasilense glnB mutants. The N terminus of NifA of H. seropedicae is a regulatory domain responsive to ammonium levels; therefore, the truncated protein has a constitutive activity, irrespective of the fixed nitrogen status (28).

The Escherichia coli and A. brasilense strains used in this work (Table 1) were grown in liquid LB (24) and NFbHPN (23), respectively. Nitrogenase activity was measured for A. brasilense cultures grown in 60-ml flasks containing NFbHP (10 ml) supplemented with ammonium chloride (5 mM) at 120 rpm and 30°C, as described previously (15). After 18 to 20 h of incubation (optical density at 600 nm, ~1.8) the cultures had zero ammonium and were derepressed for nitrogenase activity, which was assayed by the acetylene reduction method (26). Protein and ammonium were measured as described previously (3, 4).


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  		TABLE 1. Bacterial strains and plasmids

The A. brasilense transconjugants used in this work were obtained by conjugation with E. coli strain S17.1 containing plasmid pEMS136 (N-truncated HsNifA expressed constitutively) (28) or pJC1 (A. brasilense glnB) (30). The transconjugants of the A. brasilense wild type (FP2) and nifA (FP10) and glnB (7628) mutant strains carrying plasmid pEMS136 fixed nitrogen with specific activities similar to that of the wild type. Addition of ammonium chloride (0.2 mM) to derepressed A. brasilense FP2 pEMS136 caused complete inhibition of nitrogenase activity (Fig. 1A). The activity was fully recovered after approximately 30 min. A similar effect was observed for the strain FP10 pEMS136 (Fig. 1A). However, approximately 60% of nitrogenase activity (Fig. 1B) was still present 30 min after ammonium addition (0.2 or 1 mM) to the derepressed cultures of strain 7628 (glnB mutant) pEMS136. Anaerobic switch-off, however, was not affected (Fig. 2B).



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  		FIG. 1. GlnB is required for nitrogenase switch-off by ammonium ions and glutamine in A. brasilense. (A, B, and C) Ammonium chloride, 0.2 mM (open symbols) or 1 mM (filled symbols), was added at time zero to derepressed cultures of A. brasilense strains, and nitrogenase activity was monitored by the acetylene reduction method. (D) Glutamine, 1 mM (filled symbols), was added to derepressed cultures. (A) Strains FP2 (wild-type) pEMS136 ({Delta}N-Hs NifA) ({square}), FP10 (nifA mutant) pEMS136 ({Delta}N-HsNifA) ({cjs3573}), and 7628 (glnB mutant) pJC1 (A. brasilense glnB) ({triangleup}). (B) Strains FP2 pEMS136 ({blacksquare}), 7628 (glnB mutant) pEMS136 ({Delta}N-HsNifA) ({triangledown}), and 7628 pEMS136 ({blacktriangledown}). (C) Strains FP2 pEMS136 ({blacksquare}) and 2812 (glnB glnZ) pEMS136 ({Delta}N-HsNifA) ({blacklozenge}). (D) FP2, with addition of 1 mM glutamine ({blacksquare}), and 7628 pEMS136, with addition of 1 mM glutamine ({blacktriangleup}). Values are the averages of triplicate assays, and bars indicate standard deviations. Full nitrogenase activity (100%) was approximately 20 nmol ethylene · min–1 · mg protein–1.



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  		FIG. 2. Mutation in glnB did not affect uptake of 0.2 mM NH4+ or anaerobic switch-off. (A) Ammonium uptake of derepressed cultures following addition of 0.2 mM NH4Cl. Strains FP2 pEMS136 ({square}), 7628 (glnB) ({circ}), 7628 pJC1 ({triangleup}), 2812 (glnB glnZ) ({diamond}), and 7628 pEMS136 ({triangledown}). (B) Derepressed cultures of A. brasilense FP2 pEMS136 (squares), 7628 pEMS136 (inverted triangles), and 2812pEMS136 (diamonds) were maintained stationary for 10 min (filled symbols) or under aerobic conditions (open symbols); at 10 min, the cultures were returned to aeration. Full nitrogenase activity (100%) was approximately 20 nmol ethylene · min–1 · mg protein–1.

Since in A. brasilense the glnB and glnA genes are in an operon regulated by promoters located upstream from glnB (glnBp1-{sigma}70, and glnBp2-{sigma}N) and in the glnBA intergenic region (glnAp), the transposon inactivation of glnB in mutant 7628 affects the expression of glnA (6, 11). Therefore, the lack of ammonium-dependent nitrogenase inhibition could be due to the lower glutamine synthetase activity in this strain. To test if glnA was involved in the observed effect, plasmid pJC1 containing the whole of A. brasilense glnB was introduced into A. brasilense 7628. This restored full NH4+-dependent (0.2 mM) inhibition and about 90% recovery of nitrogenase activity compared to that of the control strain FP2 (wild type) or FP10 (nifA mutant) containing the pEMS136 plasmid (Fig. 1A). Furthermore, addition of 1 mM glutamine completely inactivated the nitrogenase of the wild-type FP2 strain but not that of strain 7628 pEMS136 (Fig. 1D), confirming that the failure to inhibit the nitrogenase is not due to lower production of glutamine in this strain.

Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) previously showed Fe protein ADP-ribosylation in whole cultures of A. brasilense (10). Immunodetection analysis of the Fe protein confirmed partial ADP-ribosylation of the Fe protein of A. brasilense strain 7628 pEMS136 (Fig. 3). Twenty minutes after addition of 1 mM ammonium chloride to the derepressed cultures of this strain, only 25% (50% of one subunit in each dimer) of the Fe protein was ADP-ribosylated, which is in agreement with the level of nitrogenase activity at the same point (60% [Fig. 1B]). In contrast, 48 to 50% (100% of one subunit in each dimer) of the Fe protein was ADP-ribosylated after addition of 1 mM of NH4Cl to the wild-type FP2 or FP2 pEMS136 (Fig. 3), correlating with complete inhibition of nitrogenase activity (Fig. 1B).



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  		FIG. 3. Ammonium-dependent ADP-ribosylation of the Fe protein in A. brasilense strains. Ammonium chloride (1 mM) was added to nitrogenase-derepressed cultures of the indicated strains of A. brasilense. After 20 min, Fe protein forms were identified by SDS-PAGE followed by immunodetection (20). The relative amounts of ADP-ribosylated over unmodified Fe protein (ADP/unmodified) were determined by densitometry with a Molecular Dynamics PSI densitometer.

The strain 7628 (glnB mutant) has a low methyl ammonium uptake rate (5); therefore, a decreased rate of ammonium uptake could account for the observed failure to inhibit nitrogenase completely in this strain. However, all four strains, FP2 pEMS136, 7628 pEMS136, 7628 pJC1, and 7628, took up ammonium at similar rates (Fig. 2A), indicating that ammonium uptake in the glnB mutant was equivalent to that of the wild type under our experimental conditions. Since A. brasilense seems to have a high-affinity uptake system that transports either ammonium or methyl ammonium and a low-affinity system specific for ammonium ions (29), our observations suggest that the latter can take up ammonium efficiently under the conditions employed in this work, independently of the presence of GlnB (Fig. 2A). These results show that the GlnB protein is necessary for complete ammonium-dependent ADP-ribosylation of the Fe protein in A. brasilense. The paralog protein GlnZ (7) was shown previously to be involved in nitrogenase reactivation, possibly by regulating DRAG activity in the A. brasilense 7611 (glnZ mutant) strain (15).

Ammonium-induced switch-off was totally absent in the glnB glnZ A. brasilense double mutant (strain 2812) containing plasmid pEMS136 after the addition of either 0.2 or 1 mM ammonium chloride (Fig. 1C). SDS-PAGE of whole cells, followed by immune detection of Fe protein, confirmed the complete absence of ADP-ribosylation in this mutant (Fig. 3). The uptake of ammonium in this strain was also similar to that of the wild type under the tested conditions (Fig. 2A). The double mutant also had a normal switch-off response to anaerobiosis (Fig. 2B).

The results obtained by using single glnB or glnZ mutants indicate that their translation products operate on different signaling systems in the NH4+-dependent nitrogenase switch-off/switch-on in A. brasilense. Nevertheless, the total absence of switch-off in the double mutant suggests that the effects of GlnB and GlnZ on the ammonium-induced ADP-ribosylation of the Fe protein are partially overlapping. These results show that GlnB and GlnZ are both involved in signaling to the DRAT/DRAG system for inhibition of nitrogenase activity. Since the expression of glnZ in the glnB mutant of A. brasilense is increased twofold under nitrogen-fixing conditions (5), it is possible that the residual ADP-ribosylation in mutant 7628 (glnB mutant) is caused by cross talk between the two signaling pathways. In the double mutant, there is no GlnZ; hence, it cannot replace GlnB and, therefore, no switch-off can occur. This implies a simpler situation in the wild type, with GlnB being responsible for switch-off and GlnZ being responsible for switch-on.

Proteins of the PII family have been implicated in the reversible inactivation of nitrogenase for several organisms. The regulation of the activities of both DRAT and DRAG from R. rubrum was studied for Klebsiella pneumoniae, and it was found that both enzymes required the presence of a functional glnB gene, while glnK seemed to be involved only in the regulation of DRAG (34). Moreover, an R. rubrum glnB glnJ double mutant was unable to ADP-ribosylate the Fe protein in response to either ammonium or darkness, further emphasizing their role in the regulation of DRAT and DRAG (33, 34) and indicating that either GlnB or GlnJ was required for the proper regulation of the ADP-ribosylation system in this organism (33, 35). In Methanococcus maripaludis, ammonium-dependent nitrogenase switch-off also requires at least one of two GlnB homologues, named NifI1 and NifI2, but in this case ADP-ribosylation was not detected, suggesting a different mechanism of nitrogenase switch-off/switch-on (14). In Azoarcus sp. strain BH72, ammonium-dependent ADP-ribosylation of nitrogenase required both GlnK and the high-affinity ammonium transporter AmtB (20). With R. capsulatus, recent data revealed that both GlnB and GlnK are essential for nitrogenase ADP-ribosylation (8) and that GlnB-DRAT and GlnK-DRAT interact directly (22). Our results now show that the GlnB and GlnZ proteins are required for the regulation of the DRAT and DRAG activities in A. brasilense in response to ammonium but not to anaerobiosis. Interestingly, the double mutant glnB glnJ of R. rubrum failed to ADP-ribosylate the Fe protein in response to both ammonium and darkness, indicating the involvement of different signaling pathways for energy depletion in these organisms (33, 35).

Our results also allow an explanation for the substantial nitrogenase activity in the presence of ammonium ions in A. brasilense strains FP8 and FP9 constitutively expressing K. pneumoniae NifA (23), as well as the nitrogenase activity by the FP2 strain carrying the plasmid constitutively expressing the N-terminally truncated NifA protein of H. seropedicae (pEMS136) in the presence of high concentrations of ammonium chloride (28). Under ammonium-repressing conditions, glnB is expressed at very low levels (6, 11, 18); therefore, the ADP-ribosylation system cannot be completely activated (Table 2). A. brasilense FP2 pEMS136 derepressed in the presence of a high ammonium concentration (13.4 mM of residual NH4Cl) had ~55% of nitrogenase activity (Table 2) present in cells depleted of ammonium chloride in the medium, correlating with the amount of ADP-ribosylated Fe protein (30% or 60% of one subunit in each dimer) present under the same conditions.


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  		TABLE 2. Nitrogenase activity and ammonium-dependent ADP-ribosylation of A. brasilense strains grown in the presence of NH4Cla

In conclusion, we show that the ammonium signal transducer proteins GlnB and GlnZ are necessary for reversible inactivation of nitrogenase by ammonium ions in A. brasilense but do not influence anaerobically triggered ADP-ribosylation. The different patterns of response to ammonium in these mutants suggest that the GlnB protein senses the rapid increase of ammonium concentration, whereas GlnZ is mainly responsible for signaling the return to ammonium-depleted conditions.


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ACKNOWLEDGMENTS
 
We thank C. Elmerich for providing A. brasilense strains 7628 and 2812 and Paul Ludden for Fe protein antibodies. We also thank Roseli Prado, Valter A. de Baura, and Julieta Pie for technical assistance.

We thank UFPR/TN/CNPq-Apoio ao Recém-Doutor/MCT/Pronex and Paraná Tecnologia for financial support.


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FOOTNOTES
 
* Corresponding author. Mailing address: Universidade Federal do Paraná, Departamento de Bioquímica e Biologia Molecular, Caixa Postal 19046 CEP-81531-990, Curitiba, Paraná, Brasil. Phone: 55 41 366 4398. Fax: 55 41 266 2042. E-mail for F. O. Pedrosa: fpedrosa{at}ufpr.br. E-mail for E. M. Souza: souzaem{at}ufpr.br. Back


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REFERENCES
 
    1
  1. Arcondéguy, T., R. Jack, and M. Merrick. 2001. PII signal transduction proteins, pivotal players in microbial nitrogen control. Microbiol. Mol. Biol. Rev. 65:80-105.[Abstract/Free Full Text]
    2. 2
  2. Arsene, F., P. A. Kaminski, and C. Elmerich. 1996. Modulation of NifA activity by PII in Azospirillum brasilense: evidence for a regulatory role of the NifA N-terminal domain. J. Bacteriol. 178:4830-4838.[Abstract/Free Full Text]
    3. 3
  3. Bradford, M. M. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72:248-254.[CrossRef][Medline]
    4. 4
  4. Chaney, A. L., and E. D. Marbach. 1962. Modified reagents for determination of urea and ammonia. Clin. Med. 8:130-132.
    5. 5
  5. de Zamaroczy, M. 1998. Structural homologues PII and PZ of Azospirillum brasilense provide intracellular signalling for selective regulation of various nitrogen-dependent functions. Mol. Microbiol. 29:449-463.[CrossRef][Medline]
    6. 6
  6. de Zamaroczy, M., A. Paquelin, and C. Elmerich. 1993. Functional organization of the glnB-glnA cluster of Azospirillum brasilense. J. Bacteriol. 175:2507-2515.[Abstract/Free Full Text]
    7. 7
  7. de Zamaroczy, M., A. Paquelin, G. Peltre, K. Forchhammer, and C. Elmerich. 1996. Coexistence of two structurally similar but functionally different PII proteins in Azospirillum brasilense. J. Bacteriol. 178:4143-4149.[Abstract/Free Full Text]
    8. 8
  8. Drepper, T., S. Groß, and A. F. Yakunin. 2003. Role of GlnB and GlnK in ammonium control of both nitrogenase systems in the phototrophic bacterium Rhodobacter capsulatus. Microbiology 149:2203-2212.[Abstract/Free Full Text]
    9. 9
  9. Fu, H. A., A. Hartmann, R. G. Lowery, W. P. Fitzmaurice, G. P. Roberts, and R. H. Burris. 1989. Posttranslational regulatory system for nitrogenase activity in Azospirillum spp. J. Bacteriol. 171:4679-4685.[Abstract/Free Full Text]
    10. 10
  10. Hartmann, A., H. Fu, and R. H. Burris. 1986. Regulation of nitrogenase activity by ammonium chloride in Azospirillum spp. J. Bacteriol. 165:864-870.[Abstract/Free Full Text]
    11. 11
  11. Huergo, L. F., E. M. Souza, M. B. R. Steffens, M. G. Yates, F. O. Pedrosa, and L. S. Chubatsu. 2003. Regulation of glnB gene promoter expression in Azospirillum brasilense by the NtrC protein. FEMS Microbiol. Lett. 223:33-40.[CrossRef][Medline]
    12. 12
  12. Jouanneau, Y., C. M. Meyer, and P. M. Vignais. 1983. Regulation of nitrogenase activity through iron protein interconversion into an active and inactive form in Rhodopseudomonas capsulata. Biochim. Biophys. Acta 749:318-328.
    13. 13
  13. Kanemoto, R. H., and P. W. Ludden. 1984. Effect of ammonia, darkness, and phenazine methosulfate on whole-cell nitrogenase activity and Fe protein modification in Rhodospirillum rubrum. J. Bacteriol. 158:713-720.[Abstract/Free Full Text]
    14. 14
  14. Kessler, P. S., C. Daniel, and J. A. Leigh. 2001. Ammonia switch-off of nitrogen fixation in the methanogenic archaeon Methanococcus maripaludis: mechanistic features and requirement for the novel GlnB homologues, NifI1 and NifI2. J. Bacteriol. 183:882-889.[Abstract/Free Full Text]
    15. 15
  15. Klassen, G., E. M. de Souza, M. G. Yates, L. U. Rigo, J. Inaba, and F. O. Pedrosa. 2001. Control of nitrogenase reactivation by the GlnZ protein in Azospirillum brasilense. J. Bacteriol. 183:6710-6713.[Abstract/Free Full Text]
    16. 16
  16. Kranz, R. G., and D. Foster-Hartnett. 1990. Transcriptional regulatory cascade of nitrogen fixation genes in anoxygenic photosynthetic bacteria: oxygen and nitrogen responsive factors. Mol. Microbiol. 4:1783-1800.
    17. 17
  17. Liang, Y. Y., M. de Zamaroczy, F. Arsene, A. Paquelin, and C. Elmerich. 1992. Regulation of nitrogen fixation in Azospirillum brasilense Sp7: involvement of nifA, glnA and glnB gene products. FEMS Microbiol. Lett. 79:113-119.[Medline]
    18. 18
  18. Liang, Y. Y., P. A. Kaminski, and C. Elmerich. 1991. Identification of a nifA-like regulatory gene of Azospirillum brasilense Sp7 expressed under conditions of nitrogen fixation and in the presence of air and ammonia. Mol. Microbiol. 5:2735-2744.[CrossRef][Medline]
    19. 19
  19. Lowery, R. G., L. L. Saari, and P. W. Ludden. 1986. Reversible regulation of the nitrogenase iron protein from Rhodospirillum rubrum by ADP-ribosylation in vitro. J. Bacteriol. 166:513-518.[Abstract/Free Full Text]
    20. 20
  20. Martin, D. E., and B. Reinhold-Hurek. 2002. Distinct roles of PII-like signal transmitter proteins and amtB in regulation of nif gene expression, nitrogenase activity, and posttranslational modification of NifH in Azoarcus sp. strain BH72. J. Bacteriol. 184:2251-2259.[Abstract/Free Full Text]
    21. 21
  21. Munoz-Centeno, M. C., M. T. Paneque, and F. J. Cejudo. 1997. Posttranslational regulation of nitrogenase activity by fixed nitrogen in Azotobacter chroococcum. Biochim. Biophys. Acta 1271:67-74.
    22. 22
  22. Pawlowski, A., K.-U. Riedel, W. Klipp, P. Dreiskemper, S. Groß, H. Bierhoff, T. Drepper, and B. Masepohl. 2003. Yeast two-hybrid studies on interaction of proteins involved in regulation of nitrogen fixation in the phototrophic bacterium Rhodobacter capsulatus. J. Bacteriol. 185:5240-5247.[Abstract/Free Full Text]
    23. 23
  23. Pedrosa, F. O., and M. G. Yates. 1984. Regulation of nitrogen fixation (nif) genes of Azospirillum brasilense by nif and ntr(gln) type gene products. FEMS Microbiol. Lett. 23:95-101.
    24. 24
  24. Sambrook, J., E. F. Fritsch, and T. Maniatis. 1989. Molecular cloning: a laboratory manual, 2nd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.
    25. 25
  25. Schick, H. J. 1971. Substrate and light dependent fixation of molecular nitrogen in Rhodospirillum rubrum. Arch. Microbiol. 75:89-101.
    26. 26
  26. Scholhorn, R., and R. H. Burris. 1967. Acetylene as a competitive inhibitor of N2 fixation. Proc. Natl. Acad. Sci. USA 58:213-216.[Free Full Text]
    27. 27
  27. Simon, R., E. F. Priefer, and A. Puhler. 1983. A broad host range mobilization system for ‘in vivo’ genetic engineering transposon mutagenesis in Gram negative bacteria. Bio/Technology 1:784-791.[CrossRef]
    28. 28
  28. Souza, E. M., F. O. Pedrosa, M. Drummond, L. U. Rigo, and M. G. Yates. 1999. Control of Herbaspirillum seropedicae NifA activity by ammonium ions and oxygen. J. Bacteriol. 181:681-684.[Abstract/Free Full Text]
    29. 29
  29. Van Dommelen, A., V. Keijers, J. Vanderleyden, and M. de Zamaroczy. 1998. (Methyl)ammonium transport in the nitrogen-fixing bacterium Azospirillum brasilense. J. Bacteriol. 180:2652-2659.[Abstract/Free Full Text]
    30. 30
  30. Vitorino, J. C., M. B. R. Steffens, H. B. Machado, M. G. Yates, E. M. Souza, and F. O. Pedrosa. 2001. Potential roles for the glnB and ntrYX genes in Azospirillum brasilense. FEMS Microbiol. Lett. 201:199-204.[CrossRef][Medline]
    31. 31
  31. Yakunin, A. F., and P. C. Hallenbeck. 2002. AmtB is necessary for NH4+-induced nitrogenase switch-off and ADP-ribosylation in Rhodobacter capsulatus. J. Bacteriol. 184:4081-4088.[Abstract/Free Full Text]
    32. 32
  32. Zhang, Y., A. D. Cummings, R. H. Burris, P. W. Ludden, and G. P. Roberts. 1995. Effect of ntrBC mutation on the posttranslational regulation of nitrogenase activity in Rhodospirillum rubrum. J. Bacteriol. 177:5322-5326.[Abstract/Free Full Text]
    33. 33
  33. Zhang, Y., E. L. Pohlmann, P. W. Ludden, and G. P. Roberts. 2001. Functional characterization of three GlnB homologs in the photosynthetic bacterium Rhodospirillum rubrum: roles in sensing ammonium and energy status. J. Bacteriol. 183:6159-6168.[Abstract/Free Full Text]
    34. 34
  34. Zhang, Y., E. L. Pohlmann, C. M. Halbleib, P. W. Ludden, and G. P. Roberts. 2001. Effect of PII and its homolog GlnK on reversible ADP-ribosylation of dinitrogenase reductase by heterologous expression of the Rhodospirillum rubrum dinitrogenase reductase ADP-ribosyl transferase-dinitrogenase reductase-activating glycohydrolase regulatory system in Klebsiella pneumoniae. J. Bacteriol. 183:1610-1620.[Abstract/Free Full Text]
    35. 35
  35. Zhang, Y., E. L. Pohlmann, C. M. Halbleib, P. W. Ludden, and G. P. Roberts. 2003. Regulation of nitrogen fixation by multiple PII homologs in the photosynthetic bacterium Rhodospirillum rubrum. Symbiosis 35:85-100.
    36. 36
  36. Zhang, Y., R. H. Burris, and G. P. Roberts. 1992. Cloning, sequencing, mutagenesis, and functional characterization of draT and draG genes from Azospirillum brasilense. J. Bacteriol. 174:3364-3369.[Abstract/Free Full Text]
    37. 37
  37. Zhang, Y., R. H. Burris, P. W. Ludden, and G. P. Roberts. 1994. Posttranslational regulation of nitrogenase activity in Azospirillum brasilense ntrBC mutants: ammonium and anaerobic switch-off occurs through independent signal transduction pathways. J. Bacteriol. 176:5780-5787.[Abstract/Free Full Text]
    38. 38
  38. Zhang, Y., R. H. Burris, P. W. Ludden, and G. P. Roberts. 1997. Regulation of nitrogen fixation in Azospirillum brasilense. FEMS Microbiol. Lett. 152:195-204.[CrossRef][Medline]
    39. 39
  39. Zhang, Y., R. H. Burris, P. W. Ludden, and G. P. Roberts. 1993. Posttranslational regulation of nitrogenase activity by anaerobiosis and ammonium in Azospirillum brasilense. J. Bacteriol. 175:6781-6788.[Abstract/Free Full Text]
    40. 40
  40. Zumft, W. G., and F. Castillo. 1978. Regulatory properties of the nitrogenase from Rhodopseudomonas palustris. Arch. Microbiol. 117:53-60.[CrossRef][Medline]

Applied and Environmental Microbiology, September 2005, p. 5637-5641, Vol. 71, No. 9
0099-2240/05/$08.00+0     doi:10.1128/AEM.71.9.5637-5641.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.



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