Fine tuning the specificity of the periplasmic phosphate transport receptor. Site-directed mutagenesis, ligand binding, and crystallographic studies.

Wang, Z.; Choudhary, Abha; Ledvina, P.S.; Quiocho, Florante A.

doi:10.1016/s0021-9258(17)31503-x

Cited by 58 publications

(20 citation statements)

References 27 publications

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“…This mutation in PBP had previously been shown to have a pH dependent affinity. 33 This was also shown for the coumarin-labeled PBP, but the fluorescence response is reduced and pH dependent (Fig. 2).…”

Section: Phosphate Biosensorsupporting

confidence: 64%

Development of fluorescent biosensors for probing the function of motor proteins

Webb¹

2007

Mol. BioSyst.

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Biosensors are becoming widely used both in basic research and in screening assays and reagentless sensors with fluorescent reporter groups attached to proteins form one class. This article describes the development of sensors for two small molecules, driven in particular by the need for high sensitivity and time resolution to probe mechanistic aspects of ATP-coupled motor proteins. The biosensors are for the products of the ATPase reaction, ADP and inorganic phosphate. The interplay between the possibilities for design and understanding the mechanism of the signal are discussed. Examples are described of how these sensors have been applied to understanding myosin and helicase motors.

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Section: Phosphate Biosensorsupporting

confidence: 64%

Development of fluorescent biosensors for probing the function of motor proteins

Webb¹

2007

Mol. BioSyst.

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“…This matches the observation that the binding affinity decreases with increasing ionic strength. 20 In acidic buffer solution, neither 1H nor 2H phosphate is bound to Na + , so the buffer solution does not affect K D app . Using the binding free energies of PBPs and buffer agents with phosphate and the concentration of the buffer solution, we computed the apparent binding free energies under the experimental conditions (Table 3, eq 1).…”

Section: ■ Results and Discussionmentioning

confidence: 98%

Elucidating the Phosphate Binding Mode of Phosphate-Binding Protein: The Critical Effect of Buffer Solution

Jing

Liu

et al. 2018

J. Phys. Chem. B

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Phosphate is an essential component of cell functions, and the specific transport of phosphorus into a cell is mediated by phosphate-binding protein (PBP). The mechanism of PBP-phosphate recognition remains controversial: on the basis of similar binding affinities at acidic and basic pHs, it is believed that the hydrogen network in the binding site is flexible to adapt to different protonation states of phosphates. However, only hydrogen (1H) phosphate was observed in the sub-angstrom X-ray structures. To address this inconsistency, we performed molecular dynamics simulations using the AMOEBA polarizable force field. Structural and free energy data from simulations suggested that 1H phosphate was the preferred bound form at both pHs. The binding of dihydrogen (2H) phosphate disrupted the hydrogen-bond network in the PBP pocket, and the computed affinity was much weaker than that of 1H phosphate. Furthermore, we showed that the discrepancy in the studies described above is resolved if the interaction between phosphate and the buffer agent is taken into account. The calculated apparent binding affinities are in excellent agreement with experimental measurements. Our results suggest the high specificity of PBP for 1H phosphate and highlight the importance of the buffer solution for the binding of highly charged ligands.

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“…It is worth emphasizing that, in biological systems, the specialized proteins employ a similar binding approach to binding phosphates with high selectivity [5,16,87]. However, in the development of the synthetic molecular receptors for H 2 PO 4 − , this approach has been rarely employed [88][89][90][91][92][93][94].…”

Section: Resultsmentioning

confidence: 99%

Anion Recognition by a Pincer-Type Host Constructed from Two Polyamide Macrocyclic Frameworks Jointed by a Photo-Addressable Azobenzene Switch

et al. 2022

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A sterically crowded light-responsive host 1 was synthetized with a 93% yield by applying a post-functionalization protocol utilizing the double amidation of 4,4′-azodibenzoyl dichloride with a readily available 26-membered macrocyclic amine. X-ray structures of two hydrates of trans-1 demonstrate a very different alignment of the azobenzene linkage, which is involved in T-shape or parallel-displaced π⋯π stacking interactions with the pyridine-2,6-dicarboxamide moieties from the macrocyclic backbone. Despite the rigidity of the macrocyclic framework, which generates a large steric hindrance around the azobenzene chromophore, the host 1 retains the ability to undergo a reversible cis⟷trans isomerization upon irradiation with UVA (368 nm) and blue (410 nm) light. Moreover, thermal cis→trans back-isomerization (ΔG0 = 106.5 kJ∙mol−1, t½ = 141 h) is markedly slowed down as compared to the non-macrocyclic analog. 1H NMR titration experiments in DMSO-d6/0.5% water solution reveal that trans-1 exhibits a strong preference for dihydrogenphosphate (H2PO4−) over other anions (Cl−, MeCO2−, and PhCO2−), whereas the photogenerated metastable cis-1 shows lower affinity for the H2PO4− anion.

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Fine tuning the specificity of the periplasmic phosphate transport receptor. Site-directed mutagenesis, ligand binding, and crystallographic studies.

Cited by 58 publications

References 27 publications

Development of fluorescent biosensors for probing the function of motor proteins

Development of fluorescent biosensors for probing the function of motor proteins

Elucidating the Phosphate Binding Mode of Phosphate-Binding Protein: The Critical Effect of Buffer Solution

Anion Recognition by a Pincer-Type Host Constructed from Two Polyamide Macrocyclic Frameworks Jointed by a Photo-Addressable Azobenzene Switch

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