A Short History of the Thermodynamics of Enzyme-catalyzed Reactions

Alberty, Robert A.

doi:10.1074/jbc.x400003200

Cited by 30 publications

(26 citation statements)

References 27 publications

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“…We assumed that ADP, UDP, and GDP form complexes with MgCl 2 with similar K d values, i.e. 1 mM (18). When GDP substituted efficiently for GTP or GMP-PNP as activator of UMP kinase in the reverse reaction, we used mixtures of these three nucleotides and adjusted the concentration of MgCl 2 according to the relationship indicated under "Experimental Procedures."…”

Section: Resultsmentioning

confidence: 99%

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Regulatory Mechanisms Differ in UMP Kinases from Gram-negative and Gram-positive Bacteria

Evrin

Străuţ

Slavova‐Azmanova

et al. 2007

Journal of Biological Chemistry

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In this work, we examined the regulation by GTP and UTP of the UMP kinases from eight bacterial species. The enzyme from Gram-positive organisms exhibited cooperative kinetics with ATP as substrate. GTP decreased this cooperativity and increased the affinity for ATP. UTP had the opposite effect, as it decreased the enzyme affinity for ATP. The nucleotide analogs 5-bromo-UTP and 5-iodo-UTP were 5-10 times stronger inhibitors than the parent compound. On the other hand, UMP kinases from the Gram-negative organisms did not show cooperativity in substrate binding and catalysis. Activation by GTP resulted mainly from the reversal of inhibition caused by excess UMP, and inhibition by UTP was accompanied by a strong increase in the apparent K m for UMP. Altogether, these results indicate that, depending on the bacteria considered, GTP and UTP interact with different enzyme recognition sites. In Grampositive bacteria, GTP and UTP bind to a single site or largely overlapping sites, shifting the T % R equilibrium to either the R or T form, a scenario corresponding to almost all regulatory proteins, commonly called K systems. In Gram-negative organisms, the GTP-binding site corresponds to the unique allosteric site of the Gram-positive bacteria. In contrast, UTP interacts cooperatively with a site that overlaps the catalytic center, i.e. the UMP-binding site and part of the ATP-binding site. These characteristics make UTP an original regulator of UMP kinases from Gram-negative organisms, beyond the common scheme of allosteric control.Bacterial UMP kinases represent a particular subfamily of NMP 2 kinases (1, 2). They do not share any significant sequence homology with other known NMP kinases and exist in solution as stable hexamers. A first structural model of Escherichia coli UMP kinase (3) based on the conservation of the carbamate kinase and N-acetylglutamate kinase folds (4, 5) helped to better rationalize previous site-directed mutagenesis experiments (6). The crystal structure of E. coli UMP kinase (7) indicated a similar fold between its monomers and N-acetylglutamate kinase, a dimeric enzyme (4, 5). However, the quaternary structure assembly of these two proteins is completely different (7). Deposited crystal structures of UMP kinases from other bacteria such as Pyrococcus furiosus (8), Neisseria meningitidis (Protein Data Bank code 1YBD), Hemophilus influenzae (code 2AIF), and Streptococcus pyogenes (code 1Z9D) show threedimensional structures very similar to that of the E. coli enzyme. The residues essential for binding nucleotide substrates and catalysis are conserved among all bacterial UMP kinases ( Fig. 1) (3, 9). Consequently, the active sites of these enzymes and the phosphoryl transfer mechanisms are most probably similar.Comparison of the biochemical properties of recombinant UMP kinases from Gram-negative E. coli (1, 2) and Gram-positive Streptococcus pneumoniae (10) indicated significant differences in their kinetic properties particularly in their regulation by nucleotides. Unlike the E. coli enzyme, U...

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Section: Resultsmentioning

confidence: 99%

“…The dissociation constant (K d ) of metal⅐nucleotide complexes varies within 2 orders of magnitude from 0.1 mM for MgNTP, 1 mM for MgNDP, and 20 mM for MgNMP (18,19). On the other hand, as some nucleotides played multiple roles, this resulted in mixed kinetic effects.…”

Section: Methodsmentioning

confidence: 99%

Regulatory Mechanisms Differ in UMP Kinases from Gram-negative and Gram-positive Bacteria

Evrin

Străuţ

Slavova‐Azmanova

et al. 2007

Journal of Biological Chemistry

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“…Kinases are the enzymes most often used for the phosphorylation of saccharides (Table 13-4, entries [7][8][9][10][11][12][13][14][15][16][17][18][19]. For example, glucose-6-phosphate (ll), a useful reagent for the regeneration of nicotinamide cofactors (see Chapter 15), was prepared from glucose in a one-step reaction by phosphorylation with ATP 17' 1.…”

Section: Phosphorylations With Atp As a Cofactormentioning

confidence: 99%

Formation and Cleavage of P‐O Bonds

Whitesides¹

2002

Enzyme Catalysis in Organic Synthesis

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“…In this regard, early in*vestigators recognized that the glutamine synthetase and glutaminase reactions offered special advantages for those seeking accuracy and precision in determinations of the equilibrium constant for ATP hydrolysis. In this case, summation of these reactions yields the ATPase mass action ratio: Alberty (1968;1969) and Phillips (1969) were among the first to recognize that the ability of magnesium ion to form a higher affinity complex with the ATP-form than with HATP3-explains why the ATPase equilibrium is so strongly influenced (a) by the concentration of "free" or uncomplexed magnesium ion, (b) by the concentration of protons that alter the protonation state of the nucleotide, (c) by the ionic strength of the supporting electrolyte and buffer, and (d) by the presence of other monovalent and divalent cations. More recently, Alberty and Goldberg (1992) presented a detailed consideration of the transformed Gibbs energy as the chief criterion for chemical equilibrium at specified temperature, pressure, pH, concentration of free magnesium ion, and ionic strength.…”

Section: B Atpase Thermodynamicsmentioning

confidence: 99%

Advances in the Enzymology of Glutamine Synthesis

Purich

1998

Advances in Enzymology - And Related Areas of Molecular Biology

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A Short History of the Thermodynamics of Enzyme-catalyzed Reactions

Cited by 30 publications

References 27 publications

Regulatory Mechanisms Differ in UMP Kinases from Gram-negative and Gram-positive Bacteria

Regulatory Mechanisms Differ in UMP Kinases from Gram-negative and Gram-positive Bacteria

Formation and Cleavage of P‐O Bonds

Advances in the Enzymology of Glutamine Synthesis

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