ATP-liganded form of aspartate transcarbamoylase, the logical regulatory target for allosteric control in divergent bacterial systems

Wild, James R.; Johnson, Jason L.; Loughrey, S J

doi:10.1128/jb.170.1.446-448.1988

Cited by 15 publications

(12 citation statements)

References 18 publications

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“…In each study, the effect of the nucleotides was amplified at the lower substrate concentration. When saturating concentrations of nucleotides were mixed, as previously reported (27), CTP reversed the activation by ATP. Other nucleotides had very little effect on the ATP-induced activation of ATCase; neither UTP nor GTP counteracted the ATP effect but actually promoted slight increases in activity.…”

Section: Resultssupporting

confidence: 67%

“…Furthermore, pyrimidine nucleotide pools could be coordinated with purine nucleotide pools through the competition ofATP and CTP for the regulatory control of ATCase (11,12,14,27), resulting in an activation or inhibition of de novo pyrimidine biosynthesis. From a metabolic perspective, it would seem that the intracellular chemistry of the bacterial cell is poised to maximize these allosteric interactions since the endogenous metabolic pool for aspartate approximates the S0.5 ofATCase (5 mM) and the nucleotide pools are =0.5-1.0 mM for CTP and -2-3 mM for ATP (27) under conditions of steady-state growth in E. coli. These physiological conditions would provide the most sensitive responses for the allosterically regulated enzyme since endogenous ligand concentrations approximate the half-saturating values required for inducing homotropic and heterotropic effects in vitro.…”

Section: Discussionmentioning

confidence: 99%

“…(i) CTP only inhibits the enzyme 50-70o under various physical conditions, such as altered pH, substrate concentrations, or effector concentrations (1,2,10,14,27).…”

Section: Discussionmentioning

confidence: 99%

“…Equilibrium was ensured by evaluation after 36 and 72 hr at 40C. The purities of the radioactively labeled nucleotides were determined by HPLC analysis as described (27).…”

Section: Methodsmentioning

confidence: 99%

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In the presence of CTP, UTP becomes an allosteric inhibitor of aspartate transcarbamoylase.

Wild

Loughrey-Chen

Corder

1989

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

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125

117

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The allosteric control of aspartate transcarbamoylase (ATCase, EC 2.1.3.2) of Escherichia coli involves feedback inhibition by both CTP and UTP rather than just CTP alone. It has been known that CTP functions as a heterotropic inhibitor of catalysis; however, the inhibition by CTP alone is incomplete (50-70% at various aspartate concentrations) and there is only a partial occupancy of the allosteric binding sites by CTP at saturating concentrations. The logic of these allosteric characteristics can now be understood in that UTP is a synergistic inhibitor of ATCase in the presence of CTP even though UTP has no independent effect at pH 7.0. When saturating concentrations of CTP are present, the concentration of substrate required for half-maximal activity (So.s) of the native holoenzyme for aspartate increases from 5 to 11 mM. When CTP and UTP are both present, the aspartate requirement increases further (S0.5 = 17 mM). At aspartate concentrations <5 mM, the heterotropic inhibition of ATCase is 90-95% in the presence of both pyrimidine nucleotides. UTP does enhance the binding of CTP to the holoenzyme but the number of tight binding sites does not change (n = 3). Biol. 196, have described a structural asymmetry across the molecular two-fold axis that is consistent with these CTP/UTP interactions. The synergistic inhibition of ATCase by both CTP and UTP provides a satisfying logic for ensuring a balance of endogenous pyrimidine nucleotide pools.The aspartate transcarbamoylase (ATCase, EC 2.1.3.2) of Escherichia coli provides a classic example of an enzyme subject to allosteric control by the end product of its biosynthetic pathway (1, 2). The holoenzyme of ATCase is composed of two separable catalytic trimers (c3) and three regulatory dimers (r2) that interact through a variety of specific protein-protein interfaces (3, 4). These interactions provide for specific conformational transitions (4-7) of the holoenzyme, 2(c3):3(r2), which result in changes in the catalytic rates and ligand affinities (8-10). Upon binding of substrates and/or substrate analogues the enzyme appears to make a concerted transition from an inactive "T conformation" (condensed form of the enzyme) to an expanded, more active "R conformation" (5-7). CTP affects catalysis by increasing the concentration of aspartate required to produce maximal activity. In contrast, ATP decreases the substrate requirement without altering the Vmax of the enzyme (1, 2).Both nucleotide effectors have been shown to competitively bind at the same allosteric sites even though they induce different heterotropic effects on catalysis (11)(12)(13)(14). Other pyrimidine nucleotides, specifically UTP and TTP, do not have any effect. GTP does promote a significant inhibitory effect under some conditions (25-30%) but less than that of CTP (50-70%). 8-Bromoguanosine 5'-triphosphate, as well as gadolinium complexes of ATP, CTP, and GTP, binds to the regulatory subunits, whereas UTP does not appear to bind to the native enzyme (11-13). These allosteric controls have bee...

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Section: Resultssupporting

confidence: 67%

Section: Discussionmentioning

confidence: 99%

“…(i) CTP only inhibits the enzyme 50-70o under various physical conditions, such as altered pH, substrate concentrations, or effector concentrations (1,2,10,14,27).…”

Section: Discussionmentioning

confidence: 99%

“…Equilibrium was ensured by evaluation after 36 and 72 hr at 40C. The purities of the radioactively labeled nucleotides were determined by HPLC analysis as described (27).…”

Section: Methodsmentioning

confidence: 99%

See 2 more Smart Citations

In the presence of CTP, UTP becomes an allosteric inhibitor of aspartate transcarbamoylase.

Wild

Loughrey-Chen

Corder

1989

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

Self Cite

125

117

View full text Add to dashboard Cite

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“…With CTP present, UTP binding is enhanced and preferentially directed to the low-affinity sites. Conversely, UTP binding leads to enhanced affinity for CTP at the high-affinity sites and inhibits enzyme activity by up to 95% while CTP binding alone inhibits activity to 50 to 70% (18,105,116). ATP and CTP bind in anticonformation with negative cooperativity with respect to themselves (91).…”

Section: Allosteric Site Of E Coli Atcasementioning

confidence: 99%

Allosteric Regulation of Catalytic Activity: Escherichia coli Aspartate Transcarbamoylase versus Yeast Chorismate Mutase

Helmstaedt

Krappmann

Braus

2001

Microbiol Mol Biol Rev

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Allosteric regulation of key metabolic enzymes is a fascinating field to study the structure-function relationship of induced conformational changes of proteins. In this review we compare the principles of allosteric transitions of the complex classical model aspartate transcarbamoylase (ATCase) from Escherichia coli, consisting of 12 polypeptides, and the less complicated chorismate mutase derived from baker's yeast, which functions as a homodimer. Chorismate mutase presumably represents the minimal oligomerization state of a cooperative enzyme which still can be either activated or inhibited by different heterotropic effectors. Detailed knowledge of the number of possible quaternary states and a description of molecular triggers for conformational changes of model enzymes such as ATCase and chorismate mutase shed more and more light on allostery as an important regulatory mechanism of any living cell. The comparison of wild-type and engineered mutant enzymes reveals that current textbook models for regulation do not cover the entire picture needed to describe the function of these enzymes in detail

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Aspartate Transcarbamylase from Escherichia Coli: Activity and Regulation

Lipscomb¹

1994

Advances in Enzymology - And Related Areas of Molecular Biology

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ATP-liganded form of aspartate transcarbamoylase, the logical regulatory target for allosteric control in divergent bacterial systems

Cited by 15 publications

References 18 publications

In the presence of CTP, UTP becomes an allosteric inhibitor of aspartate transcarbamoylase.

In the presence of CTP, UTP becomes an allosteric inhibitor of aspartate transcarbamoylase.

Allosteric Regulation of Catalytic Activity: Escherichia coli Aspartate Transcarbamoylase versus Yeast Chorismate Mutase

Aspartate Transcarbamylase from Escherichia Coli: Activity and Regulation

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