Determination of the Formylglycinamide Ribonucleotide Amidotransferase Ammonia Pathway by Combining 3D-RISM Theory with Experiment

Tanwar, Ajay Singh; Sindhikara, Daniel J.; Hirata, Fumio; Anand, Ruchi

doi:10.1021/cb501015r

Cited by 12 publications

(8 citation statements)

References 37 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Whereas, preformed conduits for tunnelling ammonia are seen in some, in other cases tunnel is evident upon substrate binding [7,9,20]. Examples of triad GATs in whose structures a tunnel cannot be visualized are also present [25]. 2-amino-2-desoxyisochorismate synthase and CTP synthetase are triad GATs that show a difference in the relative orientation of the two domains upon ligand binding [20,26].…”

Section: Introductionmentioning

confidence: 99%

Helices on interdomain interface couple catalysis in the ATPPase domain with allostery inPlasmodium falciparumGMP synthetase

Shivakumaraswamy

Pandey

Ballut

et al. 2019

Preprint

View full text Add to dashboard Cite

AbstractGMP synthetase catalyzes the substitution of the C2 oxo-group of the purine base in XMP with an amino-group generating GMP, the last step in the biosynthesis of GMP. This reaction involves a series of catalytic events that include hydrolysis of Gln generating ammonia in the glutamine amidotransferase (GATase) domain, activation of XMP to adenyl-XMP intermediate in the ATP pyrophosphatase (ATPPase) domain and reaction of ammonia with the intermediate to generate GMP. Inherent to the functioning of GMP synthetases is bidirectional domain crosstalk, which leads to allosteric activation of the GATase domain by substrates binding to the ATPPase domain, synchronization of the two catalytic events and tunnelling of ammonia from the GATase to the ATPPase domain. Herein, we have taken recourse to the analysis of structures of GMP synthetases, site-directed mutagenesis and, steady-state and transient kinetic assays on the Plasmodium falciparum enzyme to decipher the molecular basis of catalysis in the ATPPase domain and domain crosstalk. The results map the residues critical for catalysis in the ATPPase domain to the helices α11 and α12 that are located at the interdomain interface, and the lid-loop that follows α11. This apart, perturbing interdomain interactions involving residues on α11 and α12 impairs GATase activation. These results imply that this arrangement of helices at the domain interface with residues that play roles in ATPPase catalysis as well as domain crosstalk enables coupling ATPPase catalysis with GATase activation. Overall, the study enhances our understanding of GMP synthetases, which are drug targets in many infectious pathogens.

show abstract

Section: Introductionmentioning

confidence: 99%

Helices on interdomain interface couple catalysis in the ATPPase domain with allostery inPlasmodium falciparumGMP synthetase

Shivakumaraswamy

Pandey

Ballut

et al. 2019

Preprint

View full text Add to dashboard Cite

show abstract

“…Whether the orientational average leads to artifacts in a narrow channel lumen has, to our knowledge, not been systematically addressed. 3D-RISM has been suggested to correctly describe solvent mixtures, but applications have been mainly restricted to solvent mixtures of water and electrolyte, ,,− for which the above-mentioned challenges might be less critical. For instance, 3D-RISM was successfully used to identify ion binding sites in potassium channels. ,− Whether 3D-RISM correctly describes the distribution of solvent mixtures of uncharged molecules inside protein channels, has remained unclear and triggered some lively discussions. ,− …”

Section: Introductionmentioning

confidence: 99%

Potential of Mean Force Calculations of Solute Permeation Across UT-B and AQP1: A Comparison between Molecular Dynamics and 3D-RISM

Ariz-Extreme

Hub

2017

J. Phys. Chem. B

View full text Add to dashboard Cite

Membrane channels facilitate the efficient and selective flux of various solutes across biological membranes. A common approach to investigate the selectivity of a channel has been the calculation of potentials of mean force (PMFs) for solute permeation across the pore. PMFs have been frequently computed from molecular dynamics (MD) simulations, yet the three-dimensional reference interaction site model (3D-RISM) has been suggested as a computationally efficient alternative to MD. Whether the two methods yield comparable PMFs for solute permeation has remained unclear. In this study, we calculated potentials of mean force for water, ammonia, urea, molecular oxygen, and methanol across the urea transporter B (UT-B) and aquaporin-1 (AQP1), using 3D-RISM, as well as using MD simulations and umbrella sampling. To allow direct comparison between the PMFs from 3D-RISM and MD, we ensure that all PMFs refer to a well-defined reference area in the bulk or, equivalently, to a well-defined density of channels in the membrane. For PMFs of water permeation, we found reasonable agreement between the two methods, with differences of ≲3 kJ mol. In contrast, we found stark discrepancies for the PMFs for all other solutes. Additional calculations confirm that discrepancies between MD and 3D-RISM are not explained by the choice for the closure relation, the definition the reaction coordinate (center of mass-based versus atomic site-based), details of the molecule force field, or fluctuations of the protein. Comparison of the PMFs suggests that 3D-RISM may underestimate effects from hydrophobic solute-channel interactions, thereby, for instance, missing the urea binding sites in UT-B. Furthermore, we speculate that the orientational averages inherent to 3D-RISM might lead to discrepancies in the narrow channel lumen. These findings suggest that current 3D-RISM solvers provide reasonable estimates for the PMF for water permeation, but that they are not suitable to study the selectivity of membrane channels with respect to uncharged nonwater solutes.

show abstract

“…To measure the leaky (unproductive) ammonia activity, previously reported method was used (27). The reaction mixture mentioned above for glutamate production, containing 1 M of the test enzyme, was supplemented with ammonia assay reagent [containing a mixture of -ketoglutarate and NADPH [reduced form of nicotinamide adenine dinucleotide phosphate (NADP + )] and with l-glutamate dehydrogenase enzyme provided in the ammonia assay kit from Sigma-Aldrich.…”

Section: Leaky Ammonia Productionmentioning

confidence: 99%

Role of allosteric switches and adaptor domains in long-distance cross-talk and transient tunnel formation

et al. 2020

Self Cite

View full text Add to dashboard Cite

Transient tunnels that assemble and disassemble to facilitate passage of unstable intermediates in enzymes containing multiple reaction centers are controlled by allosteric cues. Using the 140-kDa purine biosynthetic enzyme PurL as a model system and a combination of biochemical and x-ray crystallographic studies, we show that long-distance communication between ~25-Å distal active sites is initiated by an allosteric switch, residing in a conserved catalytic loop, adjacent to the synthetase active site. Further, combinatory experiments seeded from molecular dynamics simulations help to delineate transient states that bring out the central role of nonfunctional adaptor domains. We show that carefully orchestrated conformational changes, facilitated by interplay of dynamic interactions at the allosteric switch and adaptor-domain interface, control reactivity and concomitant formation of the ammonia tunnel. This study asserts that substrate channeling is modulated by allosteric hotspots that alter protein energy landscape, thereby allowing the protein to adopt transient conformations paramount to function.

show abstract

Determination of the Formylglycinamide Ribonucleotide Amidotransferase Ammonia Pathway by Combining 3D-RISM Theory with Experiment

Cited by 12 publications

References 37 publications

Helices on interdomain interface couple catalysis in the ATPPase domain with allostery inPlasmodium falciparumGMP synthetase

Helices on interdomain interface couple catalysis in the ATPPase domain with allostery inPlasmodium falciparumGMP synthetase

Potential of Mean Force Calculations of Solute Permeation Across UT-B and AQP1: A Comparison between Molecular Dynamics and 3D-RISM

Role of allosteric switches and adaptor domains in long-distance cross-talk and transient tunnel formation

Contact Info

Product

Resources

About