Conserved Active Site Aspartates and Domain–Domain Interactions in Regulatory Properties of the Sugar Kinase Superfamily

Pettigrew, Donald W.; Smith, Gayle B.; Thomas, Kimberly P.; Dodds, D'Nette C.

doi:10.1006/abbi.1997.0444

Cited by 28 publications

(27 citation statements)

References 37 publications

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“…A conformational change of glycerol kinase is part of the catalytic cycle, and so it was proposed that the binding of EIIA Glc could lock the protein in a closed state (355). The lower inhibition by EIIA Glc observed for some mutant GlpKs affected in the domain associated with "opening" and "closing" GlpK is consistent with this concept (655,658).…”

Section: Inhibits Transcription Induction Mechanismmentioning

confidence: 78%

How Phosphotransferase System-Related Protein Phosphorylation Regulates Carbohydrate Metabolism in Bacteria

Deutscher

Francke

Postma

2006

Microbiol Mol Biol Rev

1,188

769

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SUMMARY The phosphoenolpyruvate(PEP):carbohydrate phosphotransferase system (PTS) is found only in bacteria, where it catalyzes the transport and phosphorylation of numerous monosaccharides, disaccharides, amino sugars, polyols, and other sugar derivatives. To carry out its catalytic function in sugar transport and phosphorylation, the PTS uses PEP as an energy source and phosphoryl donor. The phosphoryl group of PEP is usually transferred via four distinct proteins (domains) to the transported sugar bound to the respective membrane component(s) (EIIC and EIID) of the PTS. The organization of the PTS as a four-step phosphoryl transfer system, in which all P derivatives exhibit similar energy (phosphorylation occurs at histidyl or cysteyl residues), is surprising, as a single protein (or domain) coupling energy transfer and sugar phosphorylation would be sufficient for PTS function. A possible explanation for the complexity of the PTS was provided by the discovery that the PTS also carries out numerous regulatory functions. Depending on their phosphorylation state, the four proteins (domains) forming the PTS phosphorylation cascade (EI, HPr, EIIA, and EIIB) can phosphorylate or interact with numerous non-PTS proteins and thereby regulate their activity. In addition, in certain bacteria, one of the PTS components (HPr) is phosphorylated by ATP at a seryl residue, which increases the complexity of PTS-mediated regulation. In this review, we try to summarize the known protein phosphorylation-related regulatory functions of the PTS. As we shall see, the PTS regulation network not only controls carbohydrate uptake and metabolism but also interferes with the utilization of nitrogen and phosphorus and the virulence of certain pathogens.

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Section: Inhibits Transcription Induction Mechanismmentioning

confidence: 78%

How Phosphotransferase System-Related Protein Phosphorylation Regulates Carbohydrate Metabolism in Bacteria

Deutscher

Francke

Postma

2006

Microbiol Mol Biol Rev

1,188

769

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“…Members of the ASKHA superfamily are known to undergo domain closure which is required for catalysis (15,28,30). The catalytically essential Phe 179 residue, which is conserved in all three kinases (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…4). The ASKHA family members glycerol kinase (10,30), hexokinase (2), and phosphoglycerate kinase (5) have a conserved glycine residue in their equivalent helices (␣3 in glycerol kinase and hexokinase, ␣7 in phosphoglycerate kinase) that is postulated to be part of the hinge at which the catalytically essential domain closure occurs (2,5,10,30). In glycerol kinase and phosphoglycerate kinase a phenylalanine immediately follows this hinge glycine, and this motif is also present in the acetate kinase and propionate kinase sequences (Fig.…”

Section: Discussionmentioning

confidence: 99%

Characterization of the Acetate Binding Pocket in theMethanosarcina thermophilaAcetate Kinase

et al. 2005

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Acetate kinase catalyzes the reversible magnesium-dependent synthesis of acetyl phosphate by transfer of the ATP ␥-phosphoryl group to acetate. Inspection of the crystal structure of the Methanosarcina thermophila enzyme containing only ADP revealed a solvent-accessible hydrophobic pocket formed by residues Val 93 , Leu 122 , Phe 179 , and Pro 232 in the active site cleft, which identified a potential acetate binding site. The hypothesis that this was a binding site was further supported by alignment of all acetate kinase sequences available from databases, which showed strict conservation of all four residues, and the recent crystal structure of the M. thermophila enzyme with acetate bound in this pocket. Replacement of each residue in the pocket produced variants with K m values for acetate that were 7-to 26-fold greater than that of the wild type, and perturbations of this binding pocket also altered the specificity for longer-chain carboxylic acids and acetyl phosphate. The kinetic analyses of variants combined with structural modeling indicated that the pocket has roles in binding the methyl group of acetate, influencing substrate specificity, and orienting the carboxyl group. The kinetic analyses also indicated that binding of acetyl phosphate is more dependent on interactions of the phosphate group with an unidentified residue than on interactions between the methyl group and the hydrophobic pocket. The analyses also indicated that Phe 179 is essential for catalysis, possibly for domain closure. Alignments of acetate kinase, propionate kinase, and butyrate kinase sequences obtained from databases suggested that these enzymes have similar catalytic mechanisms and carboxylic acid substrate binding sites.Acetate kinase catalyzes the reversible magnesium-dependent phosphorylation of acetate with ATP (equation 1) and is very important for the energy-yielding metabolism of anaerobic microbes.In most fermentative anaerobes this enzyme is responsible for production of a major portion of the ATP (reverse of equation 1). Acetate kinase functions in the energy-yielding pathway for conversion of the methyl group of acetate to methane (equation 2) in Methanosarcina species.

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“…The specific activities of glycerol kinase were determined in extracts of cells that were grown overnight in LB medium at 37°C, collected by centrifugation, and disrupted by sonication after being resuspended in 0.1 M triethanolamine-HCl, 2 mM glycerol, and 2 mM ␤-mercaptoethanol (pH 7.0). Glycerol kinase activities were determined by using an ADP-coupled continuous spectrophotometric assay at pH 7.0 and 25°C with 2 mM glycerol and 2.5 mM ATP (8). Protein concentrations were determined by using the Bio-Rad assay with bovine serum albumin as the standard.…”

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confidence: 99%

Unexpected Presence of Defective glpR Alleles in Various Strains of Escherichia coli

2001

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Conserved Active Site Aspartates and Domain–Domain Interactions in Regulatory Properties of the Sugar Kinase Superfamily

Cited by 28 publications

References 37 publications

How Phosphotransferase System-Related Protein Phosphorylation Regulates Carbohydrate Metabolism in Bacteria

How Phosphotransferase System-Related Protein Phosphorylation Regulates Carbohydrate Metabolism in Bacteria

Characterization of the Acetate Binding Pocket in theMethanosarcina thermophilaAcetate Kinase

Unexpected Presence of Defective glpR Alleles in Various Strains of Escherichia coli

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