Negative homotropic cooperativity and affinity heterogeneity: preparation of yeast glyceraldehyde-3-phosphate dehydrogenase with maximal affinity homogeneity.

Gennis, L S

doi:10.1073/pnas.73.11.3928

Cited by 13 publications

(2 citation statements)

References 26 publications

(29 reference statements)

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“…However, the finding that symmetry can be preserved even during sequential filling of binding sites indicates the applicability of the symmetry principles of the MWC model to negative cooperativity, which its authors did not anticipate. At the time of the MWC model most cases of negative cooperativity were regarded as artifacts resulting from partial protein activity, as was later verified [44] for the controversial case of apparent negative cooperativity that had led to elaboration of the sequential allostery model [45]. Since that time, however, many carefully-documented examples of negative cooperativity including ArgR establish beyond doubt that both positive and negative cooperativity are common molecular strategies that serve complementary physiological purposes.…”

Section: Resultsmentioning

confidence: 98%

Symmetric Allosteric Mechanism of Hexameric Escherichia coli Arginine Repressor Exploits Competition between L-Arginine Ligands and Resident Arginine Residues

Strawn

Melicherčík²,

Green

et al. 2010

PLoS Comput Biol

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An elegantly simple and probably ancient molecular mechanism of allostery is described for the Escherichia coli arginine repressor ArgR, the master feedback regulator of transcription in L-arginine metabolism. Molecular dynamics simulations with ArgRC, the hexameric domain that binds L-arginine with negative cooperativity, reveal that conserved arginine and aspartate residues in each ligand-binding pocket promote rotational oscillation of apoArgRC trimers by engagement and release of hydrogen-bonded salt bridges. Binding of exogenous L-arginine displaces resident arginine residues and arrests oscillation, shifting the equilibrium quaternary ensemble and promoting motions that maintain the configurational entropy of the system. A single L-arg ligand is necessary and sufficient to arrest oscillation, and enables formation of a cooperative hydrogen-bond network at the subunit interface. The results are used to construct a free-energy reaction coordinate that accounts for the negative cooperativity and distinctive thermodynamic signature of L-arginine binding detected by calorimetry. The symmetry of the hexamer is maintained as each ligand binds, despite the conceptual asymmetry of partially-liganded states. The results thus offer the first opportunity to describe in structural and thermodynamic terms the symmetric relaxed state predicted by the concerted allostery model of Monod, Wyman, and Changeux, revealing that this state is achieved by exploiting the dynamics of the assembly and the distributed nature of its cohesive free energy. The ArgR example reveals that symmetry can be maintained even when binding sites fill sequentially due to negative cooperativity, which was not anticipated by the Monod, Wyman, and Changeux model. The molecular mechanism identified here neither specifies nor requires a pathway for transmission of the allosteric signal through the protein, and it suggests the possibility that binding of free amino acids was an early innovation in the evolution of allostery.

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Section: Resultsmentioning

confidence: 98%

Symmetric Allosteric Mechanism of Hexameric Escherichia coli Arginine Repressor Exploits Competition between L-Arginine Ligands and Resident Arginine Residues

Strawn

Melicherčík²,

Green

et al. 2010

PLoS Comput Biol

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“…Implicit in the foregoing is the assumption that the enzyme preparation is homogeneous. Slade has found that heterogeneity with respect to NAD affinity exists in yeast glyceraldehyde-3-phosphate dehydrogenase which is in other respects homogeneous (Gennis, 1976). On the other hand, the reordered alkylation experiment of Stallcup & Koshland (1973a) indicates that heterogeneity does not contribute to the half-of-the-sites reactivity that they observe in this enzyme.…”

Section: Discussionmentioning

confidence: 96%

Molecular symmetry and metastable states of enzymes exhibiting half-of-the-sites reactivity

Herzfeld¹,

Ichiye²,

Jung³

1981

Biochemistry

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Half-of-the-sites reactivity in oligomeric enzymes has generally been accepted as evidence for structural asymmetry between subunits. However, we show that the symmetric two-state allosteric model [Monod, J., Wyman, J., & Changeux, J.-P. (1965) J. Mol. Biol. 12, 88-118] is quantitatively consistent with half-of-the-sites reactivity data for several hexameric and tetrameric enzymes. Specifically, the time courses for both the modification and the inactivation of glutamate dehydrogenase by glutamyl alpha-chloromethyl ketone and uridine diphosphoglucose dehydrogenase by 5-(iodoacetamidoethyl)aminonaphthalene-1-sulfonic acid are fit with just five parameters for each enzyme-modifier pair. In the case of glyceraldehyde-3-phosphate dehydrogenase, the time courses for modification of the yeast enzyme by iodoacetic acid and the rabbit-muscle enzyme by 3,3,3-trifluorobromoacetone are fit with the same model, and parameter values from these fits are used to generate theoretical inactivation curves which are found to agree well with the experimentally measured inactivation. We conclude that half-of-the-sites reactivity, if it is not an artifact of residual heterogeneity, could be a kinetic phenomenon related to metastability of partially modified states of a symmetric oligomer and that asymmetry between subunits should therefore not necessarily be inferred from such behavior. If similar metastability occurs in substrate binding, it may play a significant role in mechanism of catalysis and control. In such cases, the virtual inaccessibility of the substrate binding equilibrium would preclude conventional quasi-equilibrium models for the enzyme kinetics.

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Mass Spectrometry as a Novel Approach to Probe Cooperativity in Multimeric Enzymatic Systems

Rogniaux

Sanglier

Strupat

et al. 2001

Analytical Biochemistry

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Negative homotropic cooperativity and affinity heterogeneity: preparation of yeast glyceraldehyde-3-phosphate dehydrogenase with maximal affinity homogeneity.

Cited by 13 publications

References 26 publications

Symmetric Allosteric Mechanism of Hexameric Escherichia coli Arginine Repressor Exploits Competition between L-Arginine Ligands and Resident Arginine Residues

Symmetric Allosteric Mechanism of Hexameric Escherichia coli Arginine Repressor Exploits Competition between L-Arginine Ligands and Resident Arginine Residues

Molecular symmetry and metastable states of enzymes exhibiting half-of-the-sites reactivity

Mass Spectrometry as a Novel Approach to Probe Cooperativity in Multimeric Enzymatic Systems

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