The role of monovalent cations in the ATPase reaction of DNA gyrase

Hearnshaw, S.J.; Chung, T.T.; Stevenson, C.E.M.; Maxwell, Anthony; Lawson, David M.

doi:10.1107/s1399004715002916

Cited by 17 publications

(16 citation statements)

References 45 publications

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“…While Hsp90’s adenosine triphosphate (ATP) hydrolysis has been studied under varied potassium chloride concentrations,13 corresponding information on sodium chloride was not previously available. In particular, it was unclear whether potassium and sodium would cause a similar effect, as differing results were reported concerning other GHKL ATPases: maximal ATPase activity was previously found in the presence of sodium for MutL;14 whereas for DNA gyrase, higher activity was found in the presence of potassium 15. Further differing effects were found regarding Hsp90-client interactions.…”

Section: Introductionmentioning

confidence: 86%

Efficient use of single molecule time traces to resolve kinetic rates, models and uncertainties

Schmid

Hugel

2017

The Journal of Chemical Physics

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Single molecule time traces reveal the time evolution of unsynchronized kinetic systems. Especially single molecule Förster resonance energy transfer (smFRET) provides access to enzymatically important time scales, combined with molecular distance resolution and minimal interference with the sample. Yet the kinetic analysis of smFRET time traces is complicated by experimental shortcomings-such as photo-bleaching and noise. Here we recapitulate the fundamental limits of single molecule fluorescence that render the classic, dwell-time based kinetic analysis unsuitable. In contrast, our Single Molecule Analysis of Complex Kinetic Sequences (SMACKS) considers every data point and combines the information of many short traces in one global kinetic rate model. We demonstrate the potential of SMACKS by resolving the small kinetic effects caused by different ionic strengths in the chaperone protein Hsp90. These results show an unexpected interrelation between conformational dynamics and ATPase activity in Hsp90.

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Section: Introductionmentioning

confidence: 86%

Efficient use of single molecule time traces to resolve kinetic rates, models and uncertainties

Schmid

Hugel

2017

The Journal of Chemical Physics

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“…Structural data supports that the calcium-binding cavity near the α-phosphate presented here is conserved in the GHKL (Gyrase, Hsp90, histidine Kinase and MutL) ATPase superfamily. Hearnshaw et al observed a potassium ion occupying the same pocket with a similar octahedral coordination in DNA gyrase 27 , and they pointed out that in yeast topoisomerase II, a lysine ε-ammonium group occupies the same pocket. For Hsp90s, perhaps this pocket is responsible for the mild ATPase stimulation by monovalent-cations previously observed for yeast Hsp90 28 .…”

Section: Discussionmentioning

confidence: 96%

Modulation of mitochondrial Hsp90 (TRAP1) ATPase activity by calcium and magnesium

Elnatan

2018

Preprint

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The Hsp90 protein family are ATP-dependent molecular chaperones that maintain protein homeostasis and regulate many essential cellular processes. Higher eukaryotic cells have organelle-specific Hsp90 paralogs that are adapted to each unique sub-cellular environment. The mitochondrial Hsp90, TRAP1, supports the folding and activity of electron transport components and is increasingly being appreciated as a critical player in mitochondrial signaling. It is well known that calcium plays an important regulatory role in mitochondria and can even accumulate to much higher concentrations than in the cytoplasm. Surprisingly, we find that calcium can replace the requirement for magnesium to support TRAP1 ATPase activity. Using anomalous xray diffraction, we reveal a novel calcium-binding site within the TRAP1 nucleotide-binding pocket located near the ATP a-phosphate and completely distinct from the magnesium site adjacent to the band g-phosphates. In the presence of magnesium, ATP hydrolysis by TRAP1, as with other Hsp90s, is non-cooperative, whereas calcium binding results in cooperative ATP hydrolysis by the two protomers within the Hsp90 dimer. The structural data suggest a mechanism for the cooperative behavior. Owing to the cooperativity, at high ATP concentrations, ATPase activity is higher with calcium, whereas the converse is true at low ATP concentrations. Integrating these observations, we propose a model where the divalent cations choice can control switching between non-cooperative and cooperative TRAP1 ATPase mechanisms in response ATP concentrations. This may facilitate coordination between cellular energetics, mitochondrial signaling, and protein homeostasis via alterations in the TRAP1 ATPdriven cycle.

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“…aureus GyrB numbering) was replaced based on the X-ray structure of the N-terminal 43-kDa fragment of the E . coli DNA gyrase subunit B [ 80 ] (PDB ID: 4wub). As for topoisomerase IV, the same loop region was reconstructed based on the protein sequence downloaded from UniProt (ID: P0C1S7) using the chimera model option of Schrödinger Software’s homology model building panel.…”

Section: Methodsmentioning

confidence: 99%

“…In the design phase, ULD1 and ULD2 were docked in the ATP-binding site of S. aureus GyrB and ParE using Glide XP (extra precision) protocol as implemented in Schrödinger Software. Next, in the case of gyrase B, the highly flexible loop region (between residues 105-127, S. aureus GyrB numbering) was replaced based on the X-ray structure of the N-terminal 43-kDa fragment of the E. coli DNA gyrase subunit B [80] (PDB ID: 4wub). As for topoisomerase IV, the same loop region was reconstructed based on the protein sequence downloaded from UniProt (ID: P0C1S7) using the chimera model option of Schrödinger Software's homology model building panel.…”

Section: In Silico Binding Mode Analysismentioning

confidence: 99%

Rational design of balanced dual-targeting antibiotics with limited resistance

et al. 2020

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Antibiotics that inhibit multiple bacterial targets offer a promising therapeutic strategy against resistance evolution, but developing such antibiotics is challenging. Here we demonstrate that a rational design of balanced multitargeting antibiotics is feasible by using a medicinal chemistry workflow. The resultant lead compounds, ULD1 and ULD2, belonging to a novel chemical class, almost equipotently inhibit bacterial DNA gyrase and topoisomerase IV complexes and interact with multiple evolutionary conserved amino acids in the ATPbinding pockets of their target proteins. ULD1 and ULD2 are excellently potent against a broad range of gram-positive bacteria. Notably, the efficacy of these compounds was tested against a broad panel of multidrug-resistant Staphylococcus aureus clinical strains. Antibiotics with clinical relevance against staphylococcal infections fail to inhibit a significant fraction of these isolates, whereas both ULD1 and ULD2 inhibit all of them (minimum inhibitory concentration [MIC] �1 μg/mL). Resistance mutations against these compounds are rare, have limited impact on compound susceptibility, and substantially reduce bacterial growth. Based on their efficacy and lack of toxicity demonstrated in murine infection models, these compounds could translate into new therapies against multidrug-resistant bacterial infections.

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The role of monovalent cations in the ATPase reaction of DNA gyrase

Abstract: New structures of the N-terminal 43 kDa fragment of the E. coli DNA gyrase B subunit reveal two discrete monovalent cation-binding sites that could have functional roles.

Cited by 17 publications

References 45 publications

Efficient use of single molecule time traces to resolve kinetic rates, models and uncertainties

Efficient use of single molecule time traces to resolve kinetic rates, models and uncertainties

Modulation of mitochondrial Hsp90 (TRAP1) ATPase activity by calcium and magnesium

Rational design of balanced dual-targeting antibiotics with limited resistance

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