Regulation of methionine synthesis in
 Escherichia coli
 : Effect of
 metJ
 gene product and
 S
 -adenosylmethionine on the expression of the
 metF
 gene

Shoeman, Robert L.; Redfield, Betty; Coleman, Timothy A.; Greene, Ronald C.; Smith, Albert A.; Brot, Nathan; Weissbach, Herbert

doi:10.1073/pnas.82.11.3601

Cited by 53 publications

(39 citation statements)

References 27 publications

(26 reference statements)

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“…The active form of MetJ represses the Met biosynthesis pathway (40). GSNO exposure results in decreased MetJ TFA (Fig.…”

Section: Gsno and No Inhibit Growth Through Distinct Mechanisms-mentioning

confidence: 95%

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Determination of the Escherichia coli S-Nitrosoglutathione Response Network Using Integrated Biochemical and Systems Analysis

Jarboe¹,

Hyduke²,

Tran³

et al. 2008

Journal of Biological Chemistry

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During infection or denitrification, bacteria encounter reactive nitrogen species. Although the molecular targets of and defensive response against nitric oxide (NO) in Escherichia coli are well studied, the response elements specific to S-nitrosothiols are less clear. Previously, we employed an integrated systems biology approach to unravel the E. coli NO-response network. Here we use a similar approach to confirm that S-nitrosoglutathione (GSNO) primarily impacts the metabolic and regulatory programs of E. coli in minimal medium by reaction with homocysteine and cysteine and subsequent disruption of the methionine biosynthesis pathway. Targeting of homocysteine and cysteine results in altered regulatory activity of MetJ, MetR, and CysB, activation of the stringent response and growth inhibition. Deletion of metJ or supplementation with methionine strongly attenuated the effect of GSNO on growth and gene expression. Furthermore, GSNO inhibited the ArcAB two-component system. Consistent with the underlying nitrosative and thiol-oxidative chemistry, growth inhibition and the majority of the regulatory perturbations were dependent upon GSNO internalization by the Dpp dipeptide transporter. Contrastingly, perturbation of NsrR appeared to be a result of the submicromolar levels of NO released from GSNO and did not require GSNO internalization.Escherichia coli is a normal inhabitant of the human digestive tract but is also a causative agent of disease, including cystitis, pyelonephritis, and O157:H7-mediated hemolytic uremic syndrome (1-3). During the course of infection, E. coli is exposed to reactive nitrogen species (RNS) 5 produced by the mammalian immune system. The most important RNS are, arguably, nitric oxide (NO) and nitrosothiols (RSNO), such as S-nitrosoglutathione (GSNO) (4 -6). Mapping of the regulatory networks that govern the response to these compounds is relevant to understanding infection and in the identification of potential drug targets.NO and RSNO exhibit distinct chemical reactivities. NO reacts directly with metal centers and free radicals or mediates indirect effects by formation of other RNS in conjunction with oxygen or superoxide (7). Although RSNO can release NO via homolytic cleavage or reaction with copper (I) ions (8 -10), its primary biochemical effect is direct reaction with thiol groups through transnitrosation and S-thiolation (11). Additionally, NO can freely diffuse across membranes, whereas, GSNO susceptibility has been shown to be dependent on the Dpp dipeptide ABC transporter in Salmonella (12-14). As the first step for studying RNS challenge in E. coli, previous investigations often did not distinguish between RSNO and NO-mediated effects (15, 16) and RSNOs were sometimes used as NO donors (17). Such studies set the foundation for further distinction between RSNO and NO. A recent comparison of the E. coli response to NO and GSNO during chemostat growth confirmed the expectation that NO and GSNO mediate distinct transcriptional perturbations (18).E. coli possesses a NO-specifi...

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“…The active form of MetJ represses the Met biosynthesis pathway (40). GSNO exposure results in decreased MetJ TFA (Fig.…”

Section: Gsno and No Inhibit Growth Through Distinct Mechanisms-mentioning

confidence: 95%

“…4A). Met biosynthesis repressor MetJ requires binding of Met derivative S-adenosyl-L-methionine (AdoMet) for activity (40). CysB is a dual regulator that requires binding of Cys precursor acetylserine for activity (41).…”

Section: Gsno and No Inhibit Growth Through Distinct Mechanisms-mentioning

confidence: 99%

Determination of the Escherichia coli S-Nitrosoglutathione Response Network Using Integrated Biochemical and Systems Analysis

Jarboe¹,

Hyduke²,

Tran³

et al. 2008

Journal of Biological Chemistry

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“…The promoters of these genes contain from 2 to 5 tandem repeats of the MetJ binding site, which are called metboxes and have the consensus sequence AGACGTCT (6). When MetJ is activated by S-adenosylmethionine (SAM) it binds tightly to the metboxes and shuts down transcription (7,8). The different genes repressed by MetJ not only have a variable number of metboxes but also variable sequences.…”

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confidence: 99%

MetJ repressor interactions with DNA probed by in-cell NMR

Augustus¹,

Reardon²,

Spicer

2009

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

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Atomic level characterization of proteins and other macromolecules in the living cell is challenging. Recent advances in NMR instrumentation and methods, however, have enabled in-cell studies with prospects for multidimensional spectral characterization of individual macromolecular components. We present NMR data on the in-cell behavior of the MetJ repressor from Escherichia coli, a protein that regulates the expression of genes involved in methionine biosynthesis. NMR studies of whole cells along with corresponding studies in cell lysates and in vitro preparations of the pure protein give clear evidence for extensive nonspecific interactions with genomic DNA. These interactions can provide an efficient mechanism for searching out target sequences by reducing the dependence on 3-dimensional diffusion through the crowded cellular environment. DNA provides the track for MetJ to negotiate the obstacles inherent in cells and facilitates locating and binding specific repression sites, allowing for timely control of methionine biosynthesis.DNA-protein interactions ͉ gene regulation ͉ met repressor ͉ methionine regulon ͉ nonspecific DNA T he environment in which proteins and nucleic acids function within a cell is well known to be crowded and complex (1). Understanding the influence of the numerous large and small molecule components on the biophysical and functional properties of these macromolecules in cellular environments, however, is extremely difficult (2, 3). For example, the detailed mechanism of gene regulation is of great interest, but the study of these interactions within living cells has always been a challenge.The transcriptional repressor MetJ controls the expression in Escherichia coli of the Met regulon, which is composed of at least 12 genes scattered at 7 sites around the genome (Fig. 1). These genes code for proteins that are involved in methionine biosynthesis and transport (4, 5). The promoters of these genes contain from 2 to 5 tandem repeats of the MetJ binding site, which are called metboxes and have the consensus sequence AGACGTCT (6). When MetJ is activated by S-adenosylmethionine (SAM) it binds tightly to the metboxes and shuts down transcription (7,8). The different genes repressed by MetJ not only have a variable number of metboxes but also variable sequences. MetJ is thus capable of specific binding to several variations of the consensus sequence in vivo and fine-tuning its repression of each gene (4, 9). NMR spectroscopy has recently been shown to provide a valuable strategy for studying proteins in living cells (10-13). This in-cell spectroscopy gives atomic level fingerprints and even 3-dimensional spectral data that are used to assign resonances and characterize features of individual components in the cellular milieu (12). The method has been used to show ligand binding (14), protein-protein interactions (15) and in at least 1 case, a gain of structure presumably as a result of molecular crowding inside the cell (13). Here, we report NMR results on both in-cell and cell lysate studie...

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“…Genetic studies suggest that an effector molecule controlling in vivo gpMetJ function is S-adenosyl methionine (Ado-met) (for reviews, see references 4, 8, 27). Recently, Shoeman et al (32) demonstrated that purified gpMetJ and Ado-met blocked metF expression at the level of RNA synthesis in an uncoupled DNA-directed protein synthesizing system. In the absence of Ado-met, high concentrations of gpMetJ also reduce in vitro metF expression.…”

mentioning

confidence: 99%

“…2A and B). Ado-met is the most probable candidate for this effector molecule (32,33). Therefore, the predicted decreases in the Ado-met and methionine pools, which result from the removal of the exogenous methionine supplement (Fig.…”

mentioning

confidence: 99%

Control of metF gene expression in maxicell preparations of Escherichia coli K-12: reversible action of the metJ protein and effect of vitamin B12

Emmett¹,

Johnson²

1986

J Bacteriol

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Regulation of methionine synthesis in Escherichia coli : Effect of metJ gene product and S -adenosylmethionine on the expression of the metF gene

Cited by 53 publications

References 27 publications

Determination of the Escherichia coli S-Nitrosoglutathione Response Network Using Integrated Biochemical and Systems Analysis

Determination of the Escherichia coli S-Nitrosoglutathione Response Network Using Integrated Biochemical and Systems Analysis

MetJ repressor interactions with DNA probed by in-cell NMR

Control of metF gene expression in maxicell preparations of Escherichia coli K-12: reversible action of the metJ protein and effect of vitamin B12

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