NITROGEN ASSIMILATION AND GLOBAL Regulation in e Coli

27 Jul 2003 12:20 AR AR195-MI57-07.tex AR195-MI57-07.
sgm LaTeX2e(2002/01/18) P1: IKH

10.1146/annurev.micro.57.030502.090820
Annu. Rev. Microbiol. 2003. 57:155–76

doi: 10.1146/annurev.micro.57.030502.090820
Copyright °c 2003 by Annual Reviews. All rights reserved
First published online as a Review in Advance on May 1, 2003
NITROGEN ASSIMILATION AND GLOBAL

REGULATION IN ESCHERICHIA COLI
Larry Reitzer
Access provided by Universidad Nacional Autonoma de Mexico on 04/07/16. For personal use only.
Department of Molecular and Cell Biology, The University of Texas at Dallas,

Richardson, Texas 75080-0688; email: reitzer@utdallas.edu
Annu. Rev. Microbiol. 2003.57:155-176. Downloaded from www.annualreviews.org
Key Words Ntr response, σ 54, Lrp, ppGpp, metabolic integration

■ Abstract Nitrogen limitation in Escherichia coli controls the expression of about
100 genes of the nitrogen regulated (Ntr) response, including the ammonia-assimilating
glutamine synthetase. Low intracellular glutamine controls the Ntr response through
several regulators, whose activities are modulated by a variety of metabolites. Ntr
proteins assimilate ammonia, scavenge nitrogen-containing compounds, and appear to
integrate ammonia assimilation with other aspects of metabolism, such as polyamine
metabolism and glutamate synthesis. The leucine-responsive regulatory protein (Lrp)
controls the synthesis of glutamate synthase, which controls the Ntr response, pre-
sumably through its effect on intracellular glutamine. Some Ntr proteins inhibit the
expression of some Lrp-activated genes. Guanosine tetraphosphate appears to con-
trol Lrp synthesis. In summary, a network of interacting global regulators that senses
different aspects of metabolism integrates nitrogen assimilation with other metabolic
processes.
CONTENTS
INTRODUCTION . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 156
PHYSIOLOGICAL CONTEXT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 156
Nitrogen Sources . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 156
Ammonia Assimilation and Intracellular Nitrogen Donors . . . . . . . . . . . . . . . . . . . 156
Quantitative Nitrogen Requirements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 157
NITROGEN LIMITATION AND THE NTR RESPONSE . . . . . . . . . . . . . . . . . . . . . . 159
Proteins Required for General Nitrogen-Limited Growth . . . . . . . . . . . . . . . . . . . . . 159
Regulation of GS Activity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 160
The Ntr Regulatory Cascade . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 161
Differential Expression of Ntr Genes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 164
Functions of the Ntr Response and Nac . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 165
THE σ 54 REGULON: ITS FUNCTION AND RELATION TO THE
NTR RESPONSE . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 166
Properties of σ 54 -Dependent Genes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 166
The Functions of σ 54 -Dependent Genes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 167
METABOLIC INTEGRATION: NITROGEN ASSIMILATION AND
CARBON METABOLISM . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 167
0066-4227/03/1013-0155$14.00 155
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156 REITZER
Lrp . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 167
Cyclic AMP . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 168
EINtr -NPr-EIINtr . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 168
GUANOSINE TETRAPHOSPHATE, AMINO ACID
SYNTHESIS, AND AMMONIA ASSIMILATION . . . . . . . . . . . . . . . . . . . . . . . . . . 170
CONCLUDING REMARKS: INTERACTIONS
BETWEEN GLOBAL REGULATORS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 171
INTRODUCTION
A group of scientists, which included Lavoisier, gave the name azote (without
life) to the major inert component of air—a name that is still recognizable in many
nitrogen-containing compounds. The name nitrogen was derived from niter (potas-
sium nitrate), a not-so-inert component of gunpowder. Nitrogen is present in many
intracellular metabolites and can be assimilated from inorganic or organic sources.
Its assimilation from inorganic sources requires reduction to ammonia, followed
by incorporation into intracellular metabolites. The appropriate distribution of
nitrogen among various pathways usually involves specific or local regulatory
mechanisms, such as endproduct inhibition or endproduct-mediated transcriptional
control. However, a few global regulators control the expression of genes from sev-
eral pathways and thereby coordinate metabolism. This review focuses on nitrogen
assimilation in Escherichia coli, its regulation, and the role of global regulators
in this regulation. A major theme of this review is the integration of nitrogen
assimilation with other aspects of metabolism.
PHYSIOLOGICAL CONTEXT
Nitrogen Sources
Bacteria assimilate a variety of inorganic nitrogen sources, but Escherichia coli
assimilates only ammonia aerobically. The ability to assimilate particular organic
nitrogen sources also depends on the organism. Organic nitrogen sources are usu-
ally monomeric units of macromolecules (e.g., amino acids or nucleobases) or
compounds derived from them (e.g., agmatine or putrescine). E. coli can assimilate
nitrogen from adenine, adenosine, agmatine, L- and D-alanine, allantoin (anaero-
bically), γ -aminobutyrate, ammonia, arginine, asparagine, aspartate, cytidine, cy-
tosine, glucosamine, glutamine, glutamate, glycine, ornithine, proline, putrescine,
L- and D-serine, threonine, and a few other compounds (76). Ammonia supports
the fastest growth rate and is therefore considered the preferred nitrogen source
for E. coli.
Ammonia Assimilation and Intracellular Nitrogen Donors

The primary products of ammonia assimilation are glutamate and glutamine, the
major intracellular nitrogen donors. Glutamine provides nitrogen for purines,
REGULATION OF NITROGEN METABOLISM 157
pyrimidines, asparagine (when intracellular ammonia is low), tryptophan, his-

tidine, glucosamine, p-aminobenzoate, and arginine (via carbamoyl phosphate).
Glutamate donates nitrogen for most of the E. coli transaminases.
Glutamate dehydrogenase (GDH) or glutamine synthetase (GS) assimilates the
bulk of ammonia in E. coli (75).
α-ketoglutarate + NH3 + NADPH → glutamate + NADP+ (GDH)

glutamate + ATP + NH3 → glutamine + ADP + PO−2
4 (GS)
+
glutamine + α-ketoglutarate + NADPH → 2 glutamate + NADP (GOGAT)
If GS assimilates ammonia, then glutamate synthase (GOGAT) catalyzes gluta-

mate formation. E. coli contains both the GDH- and the GS-dependent pathways,
and they are not physiologically equivalent. The ATP-consuming GS-GOGAT
pathway in E. coli is used in energy-rich environments, whereas the GDH pathway
is employed in energy-limited (presumably nitrogen-rich) environments (39, 40).
The GS-GOGAT pathway is more appropriate for ammonia assimilation in a
nitrogen-limited environment, since GS has a much lower Km for ammonia than
GDH. Ammonia assimilation by the GS-GOGAT pathway accounts for an aston-
ishing 15% of the cell’s ATP requirement, which is calculated from the fact that
synthesis of 1 g of E. coli requires 40 mmoles of carbon, about 11 mmoles of
nitrogen, and 72 mmoles of ATP (56). [GS is highly conserved (52), which is
consistent with the possibility that it assimilates ammonia in most microbial or-
ganisms. Nonetheless, it appears that alanine dehydrogenase assimilates ammonia
in Rhizobium leguminosarum bacteroids, which subsequently secrete alanine to
the pea plant (1).]
Quantitative Nitrogen Requirements

Once ammonia is assimilated, it must be distributed in the correct ratios to a
variety of compounds. The biosynthetic requirement of each nitrogen-containing
metabolite (how much must be synthesized) can be calculated from the chemical
composition of E. coli (the amount of each component that would be obtained
from complete hydrolysis of whole cells) (Table 1). The difference between the
biosynthetic requirement and the cellular composition can be quantitatively large.
For example, 1 g of E. coli contains an estimated 250 µmoles of glutamate in
protein. However, 810 µmoles of glutamate are required for arginine, glutamine,
proline, the polyamines, and peptidoglycan synthesis. Furthermore, glutamate is
the nitrogen donor for at least 1 nitrogen for 11 amino acids and the polyamines,
which requires 7108 µmoles of glutamate. Therefore, the biosynthetic requirement
for glutamate is 8168 µmoles.
Three observations are worth noting from these calculations. First, over one third
of a cell’s nitrogen is present in guanine nucleotides (1142 µmoles of nitrogen,
or 11%), arginine (1124 µmoles, or 11%), adenine nucleotides (948 µmoles, or
9%), and lysine (652 µmoles, or 6%). Second, more carbon skeletons must be syn-
thesized for aspartate and serine than for other amino acids (Table 1, footnote d).
158 REITZER
TABLE 1 Cellular composition and biosynthetic requirements for nitrogena

C for other C skeleton µmoles
Compositionb compoundsc subtotald N donatione Totalf Ng
alanine 488 55 543 543 488

arginine 281 281 281 1124
asparagine 229 229 229 458
aspartate 229 1565 1794 979 2772 229
cysteine 87 153 240 240 87

glutamate 250 810 1060 7108 8168 250
glutamine 250 250 10226 10476 500

glycine 582 418 1000 1000 582
histidine 90 90 90 270
isoleucine 276 276 276 276
leucine 428 428 428 428
lysine 326 28 354 354 652
methionine 146 7 153 153 146
phenylalanine 176 176 176 176
proline 210 210 210 210
serine 205 1409 1614 1614 205
threonine 241 276 517 517 241
tryptophan 54 54 54 108
tyrosine 131 131 131 131
valine 402 402 402 402
AMP + dAMP 190 948
GMP + dGMP 228 1142
CMP + dCMP 151 454
UMP + dTMP 161 321
Other compoundsh 380 455
a
All units are µmoles per g dry weight of E. coli grown in glucose-ammonia minimal medium. It is assumed that glutamate
is made by the GS-glutamate synthase route.
b
Chemical composition taken from (65), which should be consulted for assumptions. For the amino acids the content from
proteins is presented.
c
This refers to syntheses that require the carbon skeleton of the amino acid, e.g., methionine synthesis requires aspartate.
d
This number is the amount of the carbon skeleton that must be synthesized.
e
After nitrogen donation it is not necessary to resynthesize the carbon skeleton, which is why this number is not contained
within the previous column. For example, 7108 µmoles of α-ketoglutarate for glutamate synthesis come from deamination
of glutamate, and only 1060 µmoles come from citric acid cycle components.
f
This column shows the biosynthetic requirement, i.e., the total µmoles of each amino acid synthesized per g E. coli.
g
This number is the number of nitrogen atoms in each compound times the amount in the composition column (footnote b).
The sum of nitrogen content is about 10,300 µmoles of nitrogen atoms per gram.
h
These include putrescine, spermidine, ethanolamine, glucosamines, and cell wall components. The amounts for each of
these components have been presented (65).
Both amino acids provide carbon and nitrogen for other amino acids and nu-
cleotides. These properties distinguish aspartate and serine from other amino acids
and might provide a metabolic basis for the specificity of the major chemotactic
receptors. Third, the biosynthetic requirement for glycine (1000 µmoles) is al-
most equal to that for C1-derivatives in purine, thymine, and methionine synthesis
(1104 µmoles). This stoichiometry necessarily couples nucleotide and amino acid
synthesis and may obviate the need for more complex regulatory mechanisms. It is
conceivable that the amazingly complex regulation directed at the glycine cleavage
enzyme (38) is sufficient for fine-tuning C1 unit synthesis.

NITROGEN LIMITATION AND THE NTR RESPONSE
Proteins Required for General Nitrogen-Limited Growth

Growth in a minimal medium with a single organic nitrogen source is slower than
that with ammonia. Such growth affects the expression of about 100 genes. These
collective changes are considered the nitrogen-regulated (Ntr) response. Loss of a
few metabolic enzymes can prevent the utilization of a variety of nitrogen sources.
This section considers those enzymes.
GLUTAMINE SYNTHETASE Nitrogen-limited growth generally requires both en-

zymes of the GS-GOGAT pathway of ammonia assimilation. GS catalyzes the
only reaction of glutamine formation, and its complete loss results in glutamine
auxotrophy (75). Basal glnA (GS-encoding) expression from the glnAp1 promoter
(which occurs in a glnG mutant) is sufficient for growth with ammonia and a
few nitrogen sources (aspartate and putrescine), but not with other single nitrogen
sources (A. Kiupakas & L. Reitzer, unpublished observation). Expression of the
glnALG (or glnA-ntrBC) operon from the glnAp2 promoter is required for optimal
GS synthesis and for high levels of two important regulators of the Ntr response,
nitrogen regulator II (NRII, also called NtrB) and nitrogen regulator I (NRI, also
called NtrC), which are required for maximal GS synthesis.
GLUTAMATE SYNTHASE The gltBD operon codes for the two subunits of GOGAT
(75). The proposals that the E. coli gltBD operon also contains gltF (23) and that
GltF regulates the Ntr response (23, 24) have not withstood close scrutiny (35, 36).
gltBD mutants are pleiotrophically defective in nitrogen source utilization and
are said to have an Asm− phenotype (76). E. coli and Klebsiella pneumoniae
mutants can still utilize a few nitrogen sources either because they can readily
generate glutamate (e.g., asparagine, aspartate, glutamate, glutamine) or because
they generate ammonia so rapidly that GDH can synthesize glutamate (e.g.,
D-serine) (93). Two different explanations can account for the Asm− phenotype
(35). The glutamine-excess hypothesis proposes that GS assimilates ammonia into
glutamine, which accumulates in the absence of GOGAT (the major glutamine-
metabolizing enzyme) and prevents Ntr gene expression. The products of Ntr genes
160 REITZER
would normally generate glutamate independent of both GOGAT and GDH (e.g.,
arginine catabolism). Alternatively, the glutamate-starvation hypothesis proposes
that glutamate starvation prevents Ntr induction (35). A mechanism by which this
occurs is not stated. A hybrid hypothesis is that glutamine accumulation prevents
Ntr gene expression until glutamate starvation stops metabolism and growth. In
any case, the arguments are complex and subject to several untested assumptions
(35). It is possible that the explanation for the Asm− phenotype involves still other
factors, such as an indirect effect of glutamate starvation.
ASNB, NADE, AND THE AMIDOTRANSFERASES Defects in several nonassimilatory

enzymes can also have pleiotropic effects in nitrogen source utilization. Enteric
bacteria have ammonia- and glutamine-dependent asparagine synthetases, AsnA
and AsnB, respectively. A K. aerogenes asnB mutant fails to grow in ammonia-
restricted nitrogen-limited environments because AsnA is insufficient for aspara-
gine synthesis (77). NAD synthetases in E. coli and other bacteria are often, but
not always, ammonia dependent (19, 82). A nadE (formerly nit) mutant with
diminished NAD synthetase fails to grow in ammonia-restricted environments
(15, 82). Glutamine-dependent amidotransferases with defective glutamine bind-
ing or amide transferase can inefficiently use ammonia as an alternative substrate
(61). Mutants with such defective amidotransferases fail to grow with organic ni-
trogen sources (L. Reitzer, unpublished observation). In summary, mutants with
altered enzymes that require a high concentration of ammonia are generally de-
fective in utilization of organic nitrogen sources.
Regulation of GS Activity
GS synthesizes glutamine and assimilates ammonia, and the ratio of these functions
depends on the environment. If GDH assimilates ammonia, then glutamine’s amide
provides 25% of cellular nitrogen and the function of GS is primarily anabolic. If
the GS-GOGAT pathway assimilates ammonia, then glutamine’s amide provides
almost 100% of cellular nitrogen and the assimilatory function is quantitatively
three times more important.
FEEDBACK INHIBITION The anabolic function of GS is controlled by cumulative

feedback inhibition by alanine, glycine, serine, adenosine monophosphate (AMP),
carbamoyl phosphate, cytidine triphosphate (CTP), glucosamine-6-phosphate, his-
tidine, and tryptophan (28). These nine compounds are competitive inhibitors that
bind to the glutamate or nucleotide substrate site (28, 57, 58). The last six require
glutamine for their synthesis and can be considered products of glutamine meta-
bolism. Although serine and glycine synthesis do not require glutamine, these
two amino acids are still excellent indicators of glutamine sufficiency. Nucleotide
synthesis accounts for 74% of the glutamine requirement (ignoring glutamate syn-
thesis), which implies that nucleotide sufficiency indicates glutamine sufficiency.
Nucleotide synthesis also accounts for 42% of the glycine biosynthetic
requirement, and for most of the consumption of C1 derivatives. Therefore, high

serine and glycine probably indicate nucleotide and glutamine sufficiency. Alanine
is more difficult to rationalize as a GS inhibitor.
ADENYLYLATION Covalent adenylylation is one mechanism that controls the as-

similatory function of GS. Each of the 12 subunits of GS can be adenylylated on a
tyrosine residue, which inactivates one subunit and enhances the susceptibility of
the other subunits to cumulative feedback inhibition (87). In other words, adeny-
lylation determines the extent to which GS is anabolic or assimilatory. Loss of the

adenylylation system is a problem during the transition from a nitrogen-limited to
an ammonia-containing (nitrogen-rich) environment, when fully induced unadeny-

lylated GS rapidly depletes intracellular glutamate (53, 54). Such a loss is not a
problem for nitrogen-sufficient cells (other mechanisms regulate GS synthesis) or
for cells in which GS is normally not adenylylated (during nitrogen limitation or
during the transition to steady-state nitrogen-limited growth).
The Ntr Regulatory Cascade

THE CENTRAL ROLE OF GLUTAMINE Nitrogen limitation increases GS specific
activity, GS synthesis, and Ntr gene expression, whereas nitrogen sufficiency de-
creases all three. Several regulators and environmental factors control this coor-
dinated response, which has been extensively reviewed (3, 11, 60, 69, 70, 75, 87).
Measurements of intracellular glutamine and studies with highly purified uridylyl-
transferase (UTase)/uridylyl-removing enzyme (UR), the first enzyme of the Ntr
regulatory cascade, suggest that the absolute concentration of glutamine controls
the regulatory cascade (44, 46, 47). High glutamine stimulates UR activity, which
removes uridine monophosphate (UMP) groups from PII-UMP and GlnK-UMP,
whereas low glutamine stimulates UTase activity, which uridylylates PII and GlnK.
Glutamine apparently binds to a single site on UTase/UR, from which it controls
both activities (46).
FUNCTIONAL REDUNDANCY: PII AND GLNK PII and GlnK have 67% amino acid
identity and similar biochemical activities (5–7, 94). The phenotypes of glnB and
glnK mutants, which lack PII and GlnK, respectively, differ only slightly from a
wild-type strain (6). In contrast, a glnB glnK double mutant has serious metabolic
problems, which result from overexpression of Ntr genes (6, 12). Furthermore,
placing glnK under the glnB promoter and vice versa do not result in appreciable
phenotypic differences (5). On the other hand, several observations suggest differ-
ent functions for PII and GlnK. First, the properties of the two purified regulators
with respect to Ntr control are not identical. Second, they are not synthesized at
the same time. Third, during nitrogen-limited growth GlnK is more abundant than
PII. Fourth, GlnK and PII form heterotrimers whose properties can differ from the
homotrimers (33, 96). Finally, GlnK can have activities that PII does not have (26).
162 REITZER
THE FUNCTION OF NTR REGULATORS IN DIFFERENT ENVIRONMENTS The

functions of these regulators must be considered in the context of nitrogen-rich
and nitrogen-limited environments and in the transitions between them (summa-
rized in Figure 1). In nitrogen-rich (i.e., ammonia-containing) minimal medium,
GS is adenylylated, glnA is expressed at a basal level, and other Ntr genes are not
expressed. PII is in excess over GlnK because PII synthesis is constitutive, while
glnK is not expressed (6, 17, 95). PII is mostly deuridylylated (33) and stimulates
the adenylylating activity of adenylyltransferase (ATase) and dephosphorylation
of NRI (via NRII), which prevents induction of Ntr genes. α-Ketoglutarate at phys-
iological concentrations binds to unmodified PII and interferes with its interaction
with ATase and NRII. Therefore, α-ketoglutarate is a signal of carbon sufficiency
and relative nitrogen limitation, but only to the extent that PII is not uridylylated
(i.e., in cells with high glutamine).
During the transition to nitrogen-limited growth PII is in excess over GlnK
and is rapidly uridylylated. PII-UMP stimulates deadenylylation of GS and fails to
interact with NRII, which results in phosphorylation of NRI (3, 69). The initial level
of NRI ∼ P is sufficient for activation of glnA, but not for other Ntr operons (4, 80).
In the absence of NRII-stimulated dephosphorylation acetyl phosphate probably
contributes to NRI phosphorylation (31).
During steady-state nitrogen-limited growth the concentrations of two impor-
tant regulators, GlnK and NRI, are higher. It can be deduced that GlnK becomes
more abundant than PII on the basis of immunological assays and the predom-
inance of GlnK homotrimers and (GlnK)2(PII)1 heterotrimers over (GlnK)1(PII)2
heterotrimers and PII homotrimers (26, 96). Both PII and GlnK are readily uridy-
lylated in vivo and in vitro, and the heterotrimers are presumably rapidly uridy-
lylated (7, 33, 94, 96). Purified GlnK-UMP slowly deadenylylates GS compared
with purified PII-UMP (96). Deadenylylation in a strain with only GlnK-UMP
might be a little slower than deadenylylation in a strain with both GlnK-UMP
and PII-UMP, although this cannot be stated with certainty because cells with
different levels of GS (because of their prior growth) were compared (94). The
rate of deadenylylation suggests that the PII subunits in purified uridylylated
heterotrimers are active (96). Because GlnK can substitute for PII and medi-
ate Ntr induction (6), it can be deduced that GlnK-UMP, like PII-UMP, fails to
interact with NRII, which results in net phosphorylation of NRI. Acetyl phos-
phate is not required for optimal glnA expression but is required for arginine
−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−→
Figure 1 The Ntr regulatory cascade: regulators and effectors. Regulation is con-
sidered for (a) steady-state growth in ammonia-containing minimal medium, (b) the
transition from nitrogen-rich to nitrogen-limited growth, (c) steady-state nitrogen-
limited growth, and (d) the transition from nitrogen-limited to nitrogen-rich growth.
Abbreviations: ATase-A, adenylylating ATase; ATase-D, deadenylylating ATase; and
αKG, α-ketoglutarate.
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164 REITZER
utilization (and therefore for the ast operon) and presumably for other Ntr genes
(31, 81).
During the transition from a nitrogen-limited to a nitrogen-replete environment
transcription of glnA, an Ntr gene, is rapidly abolished (78), and GS is rapidly
adenylylated (94). GlnK is in excess over PII. Purified GlnK-UMP is deuridyly-
lated 10 times slower than PII-UMP (7). However, because purified GlnK effec-
tively stimulates NRII-mediated dephosphorylation of NRI (7), the accumulation of
deuridylylated GlnK must be sufficient to prevent glnA expression (94). However,
it is not clear whether GlnK stimulates GS adenylylation in vivo. The activity of

purified GlnK in stimulating adenylylation is 40 times less than that of PII (7, 94).
It is more likely that GlnK-independent ATase activity is sufficient for GS adeny-
lylation (7, 33) because glutamine stimulates the adenylylating activity of ATase
and nitrogen-limited cells without either GlnK or PII can rapidly adenylylate GS
(6). In addition to these properties GlnK associates with AmtB, an ammonia trans-
porter/facilitator, and inhibits its activity (26). PII also associates with AmtB but
does not inhibit its activity (26). Therefore, ammonia transport should be impaired
during the transition to an ammonia-containing, nitrogen-rich environment. These
properties have interesting implications. If AmtB and GlnK are made stoichio-
metrically, then AmtB is in excess over PII. AmtB has the potential to sequester
PII and GlnK or to allosterically affect their activities. If so, AmtB might regulate
nitrogenase synthesis in K. pneumoniae, in which GlnK controls the activity of
NifL (37, 45).
In summary, glutamine ultimately controls the activities and interactions of the
individual regulators of the Ntr regulatory cascade. Considering the importance of
glutamine, it is probably not surprising that synthesis of the two glutamine-sensing
regulators, UTase/UR and ATase, is constitutive. In contrast, nitrogen limitation
controls the synthesis of the other regulators. Glutamine also determines whether
other signals are sensed. In a high-glutamine environment α-ketoglutarate modi-
fies the activities of unuridylylated PII and GlnK, and the products of glutamine
metabolism inhibit GS. In a low-glutamine environment acetyl phosphate con-
tributes to Ntr gene expression. Until recently, it was thought that a single protein,
UTase/UR, sensed the ratio of glutamine to α-ketoglutarate. Such regulation per-
mits the possibility of a high ratio (apparent nitrogen excess) with insufficient
glutamine for numerous glutamine-dependent enzymes (functional nitrogen de-
ficiency). Sensing the absolute concentration of glutamine avoids this problem.
Instead, the ratio of glutamine to α-ketoglutarate is important only when glu-
tamine is high and there is sufficient glutamine to drive the glutamine-dependent
reactions.
Differential Expression of Ntr Genes

SEQUENTIAL EXPRESSION Several mechanisms permit differential expression of
Ntr genes. The transition to nitrogen limitation induces Ntr genes sequentially.
The low NRI ∼ P concentration initially present during the transition to nitrogen-
limited growth is sufficient for expression of the glnA operon (80). However, a
higher NRI concentration is required for nac, glnK, the astCADBE operon, and
probably other Ntr operons (4).
NITROGEN SOURCE–DEPENDENT CONTROL Growth with different nitrogen sources

induces the glnA operon equally well (84). However, this is not the case for the
activation of astCADBE, gabDTPC, and patA/ygjG and the repression of gltBD
[(35, 50, 83); C. Pybus, B. Schneider & L. Reitzer, unpublished observations]. The
basis for this differential regulation is not known.
PREVENTION OF OVEREXPRESSION Three distinct mechanisms prevent Ntr gene

overexpression. Low NRI activates the glnAp2 promoter of the glnA operon, and
high NRI partially inhibits the activation (86). The binding of NRI to low-affinity
sites apparently mediates this modulation (8). NRI activates nac expression, and
then Nac impairs this activation (32). Finally, a glnK mutant overexpresses several
Ntr genes during nitrogen starvation, which apparently reduces viability (13). This
suggests that one function of GlnK is to prevent such overexpression.
Functions of the Ntr Response and Nac

Microarray analysis, computer analysis of σ 54-dependent genes, and other studies
suggest that the vast majority of Ntr genes have been identified and that nitrogen
limitation affects about 100 genes. This section considers the functions of these
genes. The functions of individual Ntr genes within specific physiological con-
texts (e.g., argT-dependent arginine transport within the context of other arginine
transport systems in E. coli) have been reviewed (74).
AMMONIA ASSIMILATION The central role of glutamine in regulation of the Ntr

response suggests that an important physiological function of the Ntr response is to
maintain intracellular glutamine and obviously to control ammonia assimilation.
Several regulatory mechanisms suggest the importance of this function. First, GS
is synthesized before other Ntr proteins. Second, different mechanisms prevent
overexpression of glnA and other Ntr genes. Finally, nitrogen source–dependent
regulation affects expression of Ntr genes, except for glnA.
SCAVENGING Zimmer et al. proposed that numerous Ntr transport systems scav-
enge nitrogen-containing compounds (100). Nitrogen limitation induces transport
systems for several amino acids (D-alanine, arginine, aspartate, glutamate, glu-
tamine, glycine, histidine, lysine, ornithine, and D-serine), peptides (D-alanyl-
D-alanine, dipeptides, and oligopeptides), polyamines and related compounds
(putrescine, spermidine, and γ -aminobutyrate), cytosine, and nucleosides. Scav-
enging obviously spares the need to assimilate ammonia.
METABOLIC INTEGRATION Six Ntr operons, astCADBE, codBA, ddpXABCDE,

gabDTPC, patA (ygjG), and ydcSTUVW, contain genes for catabolic enzymes. It
could be argued that these enzymes contribute to scavenging. However, this would
166 REITZER
not explain why so few catabolic enzymes are under Ntr control and why nine
of them degrade compounds that are associated with polyamine metabolism. The
products of astCADBE, gabDTPC, and patA (ygjG) degrade arginine/ornithine,
γ -aminobutyrate, putrescine, respectively [(81, 83); B. Schneider, C. Pybus &
L. Reitzer, unpublished observations). YdcW degrades γ -aminobutyraldehyde, a
product of putrescine catabolism (B. Schneider & L. Reitzer, unpublished obser-
vation). In addition to these catabolic enzymes, nitrogen limitation induces two
putrescine or polyamine transport systems, potFGHI and ydcSTUVW (100). At
least 19 Ntr genes transport putrescine, degrade putrescine, or degrade the pre-
cursors of putrescine synthesis. Polyamine levels are correlated to the growth
rate (89, 92). These Ntr genes may integrate slower nitrogen-limited growth with
polyamine content.
Metabolic integration could account for why nitrogen limitation represses both
enzymes of glutamate synthesis, the first enzyme of serine synthesis, and AsnC
(the activator of asnA) (12, 35, 64, 71). Nitrogen limitation also induces a peptidase
that degrades D-alanyl-D-alanine, a precursor for peptidoglycan (100), which may
modulate peptidoglycan synthesis. In summary, Ntr genes might integrate slower
growth with several major metabolic pathways. Such a function may account for
the lethality of Ntr gene overexpression (13).
NAC Nac is an Ntr protein that represses the enzymes of glutamate and serine
synthesis and AsnC (12, 35, 64, 71). Nac activates codBA (cytosine metabolism),
ydcSTUVW (putative putrescine transport and catabolism), nupC (nucleoside trans-
port), gabDTPC (γ -aminobutyrate transport and degradation), dppABCDF (dipep-
tide transport), and fklB-cycA (100). The products of these genes might modulate
the levels of important metabolic intermediates and, by this mechanism, integrate
nitrogen assimilation with other aspects of metabolism. If so, then such integration
is dispensable (or at least redundant) because an E. coli nac mutant does not have
a dramatic phenotype and Salmonella lacks the gene (64).
THE σ 54 REGULON: ITS FUNCTION AND

RELATION TO THE NTR RESPONSE
Properties of σ 54-Dependent Genes

Expression of many Ntr genes requires σ 54, which is the only σ factor in E. coli
that is not homologous to σ 70 (62). σ 54-dependent transcription has several distinc-
tive features (16, 74). It always requires a transcriptional activator. The activators
bind to sites that are analogous to eukaryotic enhancers. The activators hydrolyze
ATP and interact with σ 54-containing RNA polymerase. The activator-RNA poly-
merase interaction requires either a DNA-bending protein, such as integration host
factor, or a DNA curvature (20, 42). Because of the absolute requirement for an
activator, σ 54-dependent expression can be completely turned off. One advantage
of such control is the wide range of activity, from very low to very high expression
(PspA and GS, products of σ 54-dependent operons, can be a few percent of E. coli
proteins). Perhaps this dynamic range is important for proteins or pathways that
must at times consume large amounts of energy or metabolic intermediates.
The Functions of σ 54-Dependent Genes

σ 54 is widespread among bacteria and is required for a variety of functions that
are not associated with nitrogen assimilation (88). About half of the 25 known
or strongly suspected σ 54-dependent operons in E. coli do not specify proteins
of nitrogen metabolism (74). FhlA activates five of these operons, which code for
proteins associated with formate hydrogen lyase. The other σ 54-dependent operons
specify proteins for the phage shock response, zinc tolerance, two products of fatty
acid catabolism, propionate and acetoacetate, and an RNA-modifying enzyme of
unknown function. The products of these genes are associated with a variety of
environmental stresses. FhlA regulon genes are thought to contribute to pH home-
ostasis during fermentation, which potentially facilitates growth or survival in an
acidic, energy-limited, anaerobic environment (14). The phage shock proteins are
required for survival in an alkaline environment and, more generally, may maintain
the proton motive force during stress (51, 97). Zinc tolerance alleviates the stress
of high zinc. Propionate and acetoacetate are also associated with stress. FadR
participates in the universal stress response by mediating fatty acid degradation,
which would generate propionate and acetoacetate for energy (27, 30). Perhaps
the products of these genes alleviate problems associated with certain stresses
that make nitrogen assimilation difficult (74). Alternatively, expression of these
genes might somehow modulate expression of the σ 54-dependent genes of nitrogen
metabolism. A conceivable modulation mechanism is competition for σ 54, which
is not an abundant protein (48). It is also possible that these genes have no obliga-
tory association with nitrogen assimilation and that strains with certain genes with
the unique properties of σ 54-dependent promoters have a selective advantage.
METABOLIC INTEGRATION: NITROGEN ASSIMILATION

AND CARBON METABOLISM
Ammonia assimilation requires energy and intermediates from central metabolism.
Several mechanisms integrate intermediary metabolism with ammonia assimila-
tion. This section considers three mechanisms that have been identified.
Lrp
The leucine-responsive regulatory protein (Lrp) is a moderately abundant DNA-
binding protein (98), but it is not a major nucleoid-binding protein (9, 10). Lrp
has several modes of action: It can either activate or repress gene expression;
leucine can reverse, enhance, or have no effect on this regulation (68). Guano-
sine tetraphosphate (ppGpp) controls Lrp synthesis (55), which accounts for its
synthesis in amino acid–poor minimal media (25) and during the transition into
168 REITZER
stationary phase (41, 90). Lrp affects the expression of about 10% of the genes
in E. coli (43, 90). Several reviews and a recent microarray analysis consider the
broader functions of Lrp (18, 67, 68, 90).
Lrp has been implicated in several aspects of the Ntr response. Lrp activates
gltBD, which means that Lrp is required for the Ntr response (29). Lrp also controls
the metabolism of alanine, serine, and glycine, which are feedback inhibitors of GS.
Furthermore, Nac, an Ntr protein, represses two Lrp-activated operons, serA and
gltBD (12, 35). Finally, evidence pieced together from different sources suggests
that Lrp and Ntr regulators might independently activate ompF, oppABCDF, ydc-
STUVW, and yeaGH (68, 90, 100). In summary, Lrp controls Ntr gene expression,
and regulators of the Ntr response modulate the expression of some Lrp-regulated
genes.
The effects of Lrp without leucine (Figure 2, top panel) and with leucine
(Figure 2, bottom panel) must be considered separately in order to understand
Lrp’s function in ammonia assimilation. Lrp without leucine activates synthesis of
GOGAT and pyridine nucleotide transhydrogenase, which together provide two
of the three substrates for ammonia assimilation. Lrp without leucine represses
two serine deaminases and alanine dehydrogenase. The net effect would appear to
favor ammonia assimilation (and amino acid synthesis) over ammonia generation
(amino acid degradation). The addition of leucine moderately reduces GOGAT
and pyridine nucleotide transhydrogenase, but increases the amino acid degrada-
tive enzymes. To the extent that nitrogen is transferred to serine and alanine via
reversible transaminations, leucine will stimulate net amino acid catabolism. Lrp
appears to be one factor that coordinates ammonia assimilation and amino acid
catabolism in an amino acid–poor environment. This hypothesis is supported by the
observation that an lrp mutant has enhanced amino acid degradation (101). It has
been proposed that Lrp mediates the transitions between feast and famine. For the
former, amino acid catabolic functions are appropriate; while for the latter, amino
acid biosynthetic functions (including ammonia assimilation) are more appropri-
ate (18, 66). If Lrp coordinates ammonia assimilation with amino acid catabolism,
then leucine is an indicator of amino acid sufficiency. Alanine may also contribute
to this regulation because alanine also binds Lrp [(59) and references cited therein].
Cyclic AMP
Cyclic AMP bound to its receptor protein interferes with expression from the
glnAp2 promoter of the glnA operon and from other Ntr promoters (91). It is
possible that the primary effect of high cyclic AMP is at the glnALG operon and
that lowering its expression prevents synthesis of sufficient NRI to activate other
Ntr genes.
EINtr-NPr-EIINtr
EINtr (PtsP), NPr (PtsO), and EIIANtr (PtsN) are paralogs of sugar phospho-
transferase transport proteins in E. coli and other organisms, which might also

Figure 2 Lrp and nitrogen assimilation. The information is from Newman et al. (68).
The filled arrows and numbers indicate activation, whereas the open arrows and numbers
indicate repression. The top panel compares the effects on the indicated pathways or en-
zymes between strains with and without Lrp, both without added leucine. The bottom panel
compares the effects of a wild-type strain with and without leucine. SerineEXT is external
serine.
170 REITZER
contribute to coordination of carbon and nitrogen metabolism. Purified EINtr, NPr,

and EIINtr form a phosphorelay, which accepts phosphate from phosphoenolpyru-
vate (72, 73). EINtr-NPr-EIINtr phosphorylates sugars 1000 times slower than EI-
HPr-EII systems, which suggests that the former has a regulatory function, not a
transport function (73). EINtr contains a GAF domain, which regulates some σ 54-
dependent activators, and therefore suggests regulatory potential (2, 79). Further-
more, Bradyrhizobium japonicum EINtr interacts with an aspartokinase isozyme,
which also suggests regulatory potential (49). Genetic evidence suggests that EINtr
and EIINtr, but not NPr, have a regulatory function. An E. coli ptsN mutant (EIINtr
deficient) grew less well with organic nitrogen sources in carbon-limited media,
which suggests positive regulation (72). The mutant grew normally with glucose
as the carbon source and expressed at least one Ntr gene normally (72). In contrast,
a K. pneumoniae ptsN mutant had increased Ntr gene expression, which suggests
negative regulation (63). An E. coli ptsP (EINtr deficient) has not been isolated;
but loss of EINtr in other organisms prevents the expression of important genes
(49, 85). ptsO and ptsN are members of the putative rpoN-yhbH-ptsN-yhbJ-ptsO
operon, whereas ptsP is unlinked (72, 73). yhbH, yhbJ, and ptsO in E. coli, and
their homologs in P. putida, do not appear to contribute to regulation (21, 72). A
model for how the EINtr-NPr-EIINtr system might affect gene expression is not
obvious from these results, and none has been presented.
GUANOSINE TETRAPHOSPHATE, AMINO ACID

SYNTHESIS, AND AMMONIA ASSIMILATION
Slower growth and elevated ppGpp activate lrp expression, which in turn is required
for the Ntr response (29, 55). Therefore, the factors that regulate ppGpp synthesis
are important in the control of nitrogen metabolism. (Where it is stated that ppGpp
controls something, what is meant is that ppGpp appears to mediate this control.
The intent is not to exclude potential indirect effects, such as diminished GTP,
or other potential mechanisms of growth rate control.) Two factors control the
ppGpp concentration: tRNA aminoacylation and energy source availability (22). In
general, ppGpp stimulates amino acid synthesis and inhibits nucleotide synthesis
and nucleobase salvage (22). This pattern suggests a preference for amino acid
synthesis over nucleotide synthesis, which is appropriate with diminished tRNA
aminoacylation.
Nitrogen availability determines which pathways of amino acid synthesis are
affected. In a glucose-ammonia minimal medium, ppGpp0 strains, which can-
not synthesize ppGpp, absolutely require arginine, glycine, histidine, leucine,
phenylalanine, serine, threonine, and valine and have weak or strain-dependent
requirements for lysine, methionine, and tyrosine (99). These strains do not require
alanine, asparagine, aspartate, cysteine, glutamate, glutamine, or proline (99). In
a nitrogen-limited environment, ppGpp via Lrp is required for glutamate synthe-
sis, induction of the Ntr response, and ammonia assimilation, and therefore the
synthesis of all the amino acids.
CONCLUDING REMARKS: INTERACTIONS

BETWEEN GLOBAL REGULATORS
A hierarchy of global regulators affects the pathways of nitrogen metabolism.
Elevated ppGpp stimulates several pathways of amino acid synthesis and inhibits
pathways of nucleotide synthesis. ppGpp activates the synthesis of Lrp, which may
fine-tune the balance between amino acid synthesis and degradation. Lrp positively
controls the synthesis of GOGAT, which affects the level of glutamine, the most
important effector of the Ntr response. Finally, the regulators of the Ntr response
control ammonia assimilation. Numerous environmental factors modify the ac-
tivities of these global regulators. The interplay of these environmental factors is

both complex and subtle. For example, carbon limitation through growth rate con-
trol increases ppGpp, which via Lrp is required for induction of the Ntr response.
However, cyclic AMP interferes with this induction by an unknown mechanism.
Another example is the effect of impaired protein synthesis by uncharged tRNA,
which increases ppGpp and Lrp. Lrp, through its control of glutamate synthase,
induces the Ntr response, which stimulates expression of polyamine catabolic
enzymes. Diminished polyamines complete the regulatory circuit, which should
impair protein synthesis. Both of these examples show mechanisms by which
global regulators and their effectors integrate ammonia assimilation with other
aspects of metabolism.
E. coli Lrp has limited sequence similarity with homologous proteins in other
organisms and is not a global regulator in at least one organism (34). Perhaps
the entire global regulatory network that controls nitrogen metabolism is also not
conserved, but instead has evolved in enteric and related bacteria in response to
environments with large and rapid fluctuations in nutrient availability.
ACKNOWLEDGMENTS
This work was supported in part by grant MCB-0077904 from the National Science
Foundation.
The Annual Review of Microbiology is online at http://micro.annualreviews.org
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P1: FRK
July 27, 2003 15:23 Annual Reviews AR195-FM
Annual Review of Microbiology

Volume 57, 2003
CONTENTS
FRONTISPIECE, Julian Davies xii

GATHERING NO MOSS, Julian Davies 1
MOLECULAR PATHOGENICITY OF THE ORAL OPPORTUNISTIC PATHOGEN

ACTINOBACILLUS ACTINOMYCETEMCOMITANS, Brian Henderson,
Sean P. Nair, John M. Ward, and Michael Wilson 29
BRUCELLA STATIONARY-PHASE GENE EXPRESSION AND VIRULENCE,
R. Martin Roop II, Jason M. Gee, Gregory T. Robertson,
John M. Richardson, Wai-Leung Ng, and Malcolm E. Winkler 57
HOW BACTERIA ASSEMBLE FLAGELLA, Robert M. Macnab 77
A SALVAGE PATHWAY FOR PROTEIN SYNTHESIS: TMRNA AND
TRANS-TRANSLATION, Jeffrey H. Withey and David I. Friedman 101
ASSEMBLY DYNAMICS OF THE BACTERIAL CELL DIVISION PROTEIN
FTSZ: POISED AT THE EDGE OF STABILITY, Laura Romberg
and Petra Anne Levin 125
NITROGEN ASSIMILATION AND GLOBAL REGULATION IN ESCHERICHIA COLI,
Larry Reitzer 155
ON THE TRAIL OF A CEREAL KILLER: EXPLORING THE BIOLOGY OF
MAGNAPORTHE GRISEA, Nicholas J. Talbot 177
BACTERIAL MEMBRANE LIPIDS: WHERE DO WE STAND? John E. Cronan 203
SPATIAL AND TEMPORAL CONTROL OF DIFFERENTIATION AND CELL CYCLE
PROGRESSION IN CAULOBACTER CRESCENTUS, Nora Ausmees and
Christine Jacobs-Wagner 225
BACTERIAL MOTILITY ON A SURFACE: MANY WAYS TO A COMMON GOAL,
Rasika M. Harshey 249
TRANSPOSABLE ELEMENTS IN FILAMENTOUS FUNGI, Marie-Josée Daboussi
and Pierre Capy 275
BACTERIOPHAGE-INDUCED MODIFICATIONS OF HOST RNA POLYMERASE,
Sergei Nechaev and Konstantin Severinov 301
VACCINIA VIRUS MOTILITY, Geoffrey L. Smith, Brendan J. Murphy, and
Mansun Law 323
vi
P1: FRK
July 27, 2003 15:23 Annual Reviews AR195-FM
CONTENTS vii
MEASLES VIRUS 1998–2002: PROGRESS AND CONTROVERSY, Glenn F. Rall 343

THE UNCULTURED MICROBIAL MAJORITY, Michael S. Rappé and
Stephen J. Giovannoni 369
PATHWAYS OF OXIDATIVE DAMAGE, James A. Imlay 395
GENE ORGANIZATION: SELECTION, SELFISHNESS, AND SERENDIPITY,
Jeffrey G. Lawrence 419
MULTIPLE SIGMA SUBUNITS AND THE PARTITIONING OF BACTERIAL
TRANSCRIPTION SPACE, Tanja M. Gruber and Carol A. Gross 441

NATURAL SELECTION AND THE EMERGENCE OF A MUTATION PHENOTYPE:
AN UPDATE OF THE EVOLUTIONARY SYNTHESIS CONSIDERING

MECHANISMS THAT AFFECT GENOME VARIATION, Lynn Helena Caporale 467
ARCHAEAL DNA REPLICATION: EUKARYAL PROTEINS IN A BACTERIAL
CONTEXT, Beatrice Grabowski and Zvi Kelman 487
MOLECULAR GENETICS OF MYCOBACTERIUM TUBERCULOSIS PATHOGENESIS,
Josephine E. Clark-Curtiss and Shelley E. Haydel 517
THE BACTERIAL RECA PROTEIN AS A MOTOR PROTEIN, Michael M. Cox 551
DNA MISMATCH REPAIR: MOLECULAR MECHANISMS AND BIOLOGICAL
FUNCTION, Mark J. Schofield and Peggy Hsieh 579
KAPOSI’S SARCOMA–ASSOCIATED HERPESVIRUS IMMUNOEVASION AND
TUMORIGENESIS: TWO SIDES OF THE SAME COIN? Patrick S. Moore
and Yuan Chang 609
THE SECRET LIVES OF THE PATHOGENIC MYCOBACTERIA,
Christine L. Cosma, David R. Sherman, and Lalita Ramakrishnan 641
BACTERIAL BIOFILMS: AN EMERGING LINK TO DISEASE PATHOGENESIS,
Matthew R. Parsek and Pradeep K. Singh 677
INDEXES
Subject Index 703
Cumulative Index of Contributing Authors, Volumes 53–57 739
Cumulative Index of Chapter Titles, Volumes 53–57 742
ERRATA
An online log of corrections to Annual Review of Microbiology chapters
(if any, 1997 to the present) may be found at http://micro.annualreviews.org/

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Annu. Rev. Microbiol. 2003. 57:155–76

NITROGEN ASSIMILATION AND GLOBAL

Department of Molecular and Cell Biology, The University of Texas at Dallas,

Key Words Ntr response, σ 54, Lrp, ppGpp, metabolic integration

Ammonia Assimilation and Intracellular Nitrogen Donors

REGULATION OF NITROGEN METABOLISM 157

pyrimidines, asparagine (when intracellular ammonia is low), tryptophan, his-

α-ketoglutarate + NH3 + NADPH → glutamate + NADP+ (GDH)

glutamine + α-ketoglutarate + NADPH → 2 glutamate + NADP (GOGAT)

If GS assimilates ammonia, then glutamate synthase (GOGAT) catalyzes gluta-

Quantitative Nitrogen Requirements

TABLE 1 Cellular composition and biosynthetic requirements for nitrogena

alanine 488 55 543 543 488

cysteine 87 153 240 240 87

glutamine 250 250 10226 10476 500

REGULATION OF NITROGEN METABOLISM 159

enzyme (38) is sufficient for fine-tuning C1 unit synthesis.

NITROGEN LIMITATION AND THE NTR RESPONSE

Proteins Required for General Nitrogen-Limited Growth

GLUTAMINE SYNTHETASE Nitrogen-limited growth generally requires both en-

ASNB, NADE, AND THE AMIDOTRANSFERASES Defects in several nonassimilatory

FEEDBACK INHIBITION The anabolic function of GS is controlled by cumulative

REGULATION OF NITROGEN METABOLISM 161

requirement, and for most of the consumption of C1 derivatives. Therefore, high

ADENYLYLATION Covalent adenylylation is one mechanism that controls the as-

lylation determines the extent to which GS is anabolic or assimilatory. Loss of the

an ammonia-containing (nitrogen-rich) environment, when fully induced unadeny-

The Ntr Regulatory Cascade

THE FUNCTION OF NTR REGULATORS IN DIFFERENT ENVIRONMENTS The

REGULATION OF NITROGEN METABOLISM

it is not clear whether GlnK stimulates GS adenylylation in vivo. The activity of

Differential Expression of Ntr Genes

REGULATION OF NITROGEN METABOLISM 165

NITROGEN SOURCE–DEPENDENT CONTROL Growth with different nitrogen sources

PREVENTION OF OVEREXPRESSION Three distinct mechanisms prevent Ntr gene

Functions of the Ntr Response and Nac

AMMONIA ASSIMILATION The central role of glutamine in regulation of the Ntr

METABOLIC INTEGRATION Six Ntr operons, astCADBE, codBA, ddpXABCDE,

THE σ 54 REGULON: ITS FUNCTION AND

Properties of σ 54-Dependent Genes

REGULATION OF NITROGEN METABOLISM 167

The Functions of σ 54-Dependent Genes

METABOLIC INTEGRATION: NITROGEN ASSIMILATION

REGULATION OF NITROGEN METABOLISM 169

contribute to coordination of carbon and nitrogen metabolism. Purified EINtr, NPr,

GUANOSINE TETRAPHOSPHATE, AMINO ACID

REGULATION OF NITROGEN METABOLISM 171

CONCLUDING REMARKS: INTERACTIONS

tivities of these global regulators. The interplay of these environmental factors is

The Annual Review of Microbiology is online at http://micro.annualreviews.org

184:5364–75 regulation of the glnALG operon of Es-

REGULATION OF NITROGEN METABOLISM 173

Biochim. Biophys. Acta 1477:122–45 sory protein GcvR to disrupt a GcvA/

tamine synthetase adenylylation state. adenylylated glutamine synthetase from

REGULATION OF NITROGEN METABOLISM 175

Chemical composition of Escherichia mate, aspartate, asparagine, L-alanine, and

Annu. Rev. Microbiol. 49:747–75 sion of glnA in Escherichia coli is reg-

inactivation of a gene homologous to 94. van Heeswijk WC, Hoving S, Molenaar

glnA during nitrogen-limited growth. J. cascade are not regulated by nitrogen in

87. Stadtman ER. 1990. Discovery of glu- 57

Annual Review of Microbiology