•Hybrid catalysts development by incorporating a coordination complex into a biomolecule.•Upgrade of coordination catalysts upon interaction with biomolecules.•Chirality transfer from a biomolecule ...to organic substrates.•Chemogenetic optimization of hybrid catalysts.
Artificial metalloenzymes, with their high selectivity and specificity combined with a wide scope of reactivity and substrates, constitute an original approach for catalyst development. Different strategies have been proposed for their elaboration, proceeding from modification of natural enzymes using bioengineering methods to de novo protein design. Another bio-inspired methodology for the development of hybrid catalysts consists in the incorporation of coordination complexes into biomolecules, with the aim to upgrade their catalytic abilities. In these systems, the reaction performed by the naked catalyst is modulated by the well-defined structure of the host biomolecule. This conveys added value to the catalyst, such as enantioselectivity or chemoselectivity. DNA, apo-enzymes, proteins and peptides have been engaged in this approach, affording a wide diversity of reactivities and substrates. The resulting systems can then be improved by combined chemical and bioengineering optimization, allowing access to powerful catalysts. Because this approach can virtually be applied to any biomolecule or coordination complex, the elaboration of bio-based hybrid catalysts seems promising for advance in catalysis.
Full text
Available for:
GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK, ZRSKP
Recently, a super uranyl binding protein (SUP) was developed, which exhibits excellent sensitivity/selectivity to bind uranyl ions. It can be immobilized onto a surface in sensing devices to detect ...uranyl ions. Here, sum frequency generation (SFG) vibrational spectroscopy was applied to probe the interfacial structures of surface-immobilized SUP. The collected SFG spectra were compared to the calculated orientation-dependent SUP SFG spectra using a one-excitonic Hamiltonian approach based on the SUP crystal structures to deduce the most likely surface-immobilized SUP orientation(s). Furthermore, discrete molecular dynamics (DMD) simulation was applied to refine the surface-immobilized SUP conformations and orientations. The immobilized SUP structures calculated from DMD simulations confirmed the SUP orientations obtained from SFG data analyzed based on the crystal structures and were then used for a new round of SFG orientation analysis to more accurately determine the interfacial orientations and conformations of immobilized SUP before and after uranyl ion binding, providing an in-depth understanding of molecular interactions between SUP and the surface and the effect of uranyl ion binding on the SUP interfacial structures. We believe that the developed method of combining SFG measurements, DMD simulation, and Hamiltonian data analysis approach is widely applicable to study biomolecules at solid/liquid interfaces.
Full text
Available for:
IJS, KILJ, NUK, PNG, UL, UM
In various malaria-endemic regions, the appearance of resistance has precluded the use of pyrimidine-based antifolate drugs. Here, a three-step fragment screening was used to identify new ...non-pyrimidine Plasmodium falciparum dihydrofolate reductase (PfDHFR) inhibitors. Starting from a 1163-fragment commercial library, a two-step differential scanning fluorimetry screen identified 75 primary fragment hits. Subsequent enzyme inhibition assay identified 11 fragments displaying IC
50
in the 28-695 μM range and selectivity for PfDHFR. In addition to the known pyrimidine, three new anti-PfDHFR chemotypes were identified. Fragments from each chemotype were successfully co-crystallized with PfDHFR, revealing a binding in the active site, in the vicinity of catalytic residues, which was confirmed by molecular docking on all fragment hits. Finally, comparison with similar non-hit fragments provides preliminary input on available growth vectors for future drug development.
Full text
Available for:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
In the fight towards eradication of malaria, identifying compounds active against new drug targets constitutes a key approach.
7,8-dihydro-6-hydroxymethylpterin-pyrophosphokinase (
HPPK) has been ...advanced as a promising target, as being part of the parasite essential folate biosynthesis pathway while having no orthologue in the human genome. However, no drug discovery efforts have been reported on this enzyme. In this study, we conducted a three-step screening of our in-house antifolate library against
HPPK using a newly designed
HPPK-GFP protein construct. Combining virtual screening, differential scanning fluorimetry and enzymatic assay, we identified 14 compounds active against
HPPK. Compounds' binding modes were investigated by molecular docking, suggesting competitive binding with the HMDP substrate. Cytotoxicity and in vitro ADME properties of hit compounds were also assessed, showing good metabolic stability and low toxicity. The most active compounds displayed low micromolar IC
against drug-resistant parasites. The reported hit compounds constitute a good starting point for inhibitor development against
HPPK, as an alternative approach to tackle the malaria parasite.
Full text
Available for:
IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
An improved production and purification method for Alzheimer's disease related methionine-modified amyloid-β 1-40 and 1-42 peptides is proposed, taking advantage of the formation of inclusion body in ...Escherichia coli. A Thioflavin-S assay was set-up to evaluate inclusion body formation during growth and optimize culture conditions for amyloid-β peptides production. A simple and fast purification protocol including first the isolation of the inclusion bodies and second, two cycles of high pH denaturation/ neutralization combined with an ultrafiltration step on 30-kDa cut-off membrane was established. Special attention was paid to purity monitoring based on a rational combination of UV spectrophotometry and SDS-PAGE analyses at the various stages of the process. It revealed that this chromatography-free protocol affords good yield of high quality peptides in term of purity. The resulting peptides were fully characterized and are appropriate models for highly reproducible in vitro aggregation studies.
Full text
Available for:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The design of metal-binding sites in proteins that combine high affinity with high selectivity for the desired metal ion remains a challenging goal. Recently, a protein designed to display femtomolar ...affinity for
UO
2
2
+
, dubbed “Super Uranyl-binding Protein” (SUP), was described, with potential applications for removing
UO
2
2
+
in water. Although it discriminated most metal ions present in seawater, the protein showed a surprisingly high affinity for Cu
2+
ions. Here, we have investigated Cu
2+
binding to SUP using a combination of electron paramagnetic resonance, fluorescence and circular dichroism spectroscopies. Our results provide evidence for two Cu
2+
binding sites on SUP that are distinct from the
UO
2
2
+
binding site, but one of which interferes with
UO
2
2
+
binding. They further suggest that in solution the protein's secondary structure changes significantly in response to binding
UO
2
2
+
; in contrast, the crystal structures of the apo- and holo-protein are almost superimposable. These results provide insights for further improving the selectivity of SUP for
UO
2
2
+
, paving the way toward protein-based biomaterials for decontamination and/or recovery of uranium.
Enzymes immobilized on solid supports have important and industrial and medical applications. However, their uses are limited by the significant reductions in activity and stability that often ...accompany the immobilization process. Here we review recent advances in our understanding of the molecular level interactions between proteins and supporting surfaces that contribute to changes in stability and activity. This understanding has been facilitated by the application of various surface-sensitive spectroscopic techniques that allow the structure and orientation of enzymes at the solid/liquid interface to be probed, often with monolayer sensitivity. An appreciation of the molecular interactions between enzyme and surface support has allowed the surface chemistry and method of enzyme attachement to be fine-tuned such that activity and stability can be greatly enhanced. These advances suggest that a much wider variety of enzymes may eventually be amenable to immobilization as green catalysts.
Interactions between immobilized enzymes and supporting surfaces are complex and context-dependent and can significantly alter enzyme structure, stability and activity.
As pregnant women and young children remain the first victims of malaria worldwide, the search for new antimalarials has been focusing on compounds with a high safety profile and extended efficacy. ...In a previous study, a rigid biphenyl PfDHFR inhibitor was developed by fragment-based screening, displaying sub nM enzyme inhibition but poor antiparasitic activity, presumably due to its low flexibility. Here, we report a new series of compounds that combines the biphenyl fragment with a flexible linker. Interestingly, their mode of binding differs from previously reported compounds, taking advantage of strong hydrophobic interaction. The new flexible biphenyl compounds show overall improved antiparasitic activity compared to rigid ones, with the best compound displaying a 2 nM antiplasmodial IC50 and suitable drug-like properties. This confirms the importance of compound flexibility for antimalarial activity and opens the way to new opportunities for antimalarial drug design.As pregnant women and young children remain the first victims of malaria worldwide, the search for new antimalarials has been focusing on compounds with a high safety profile and extended efficacy. In a previous study, a rigid biphenyl PfDHFR inhibitor was developed by fragment-based screening, displaying sub nM enzyme inhibition but poor antiparasitic activity, presumably due to its low flexibility. Here, we report a new series of compounds that combines the biphenyl fragment with a flexible linker. Interestingly, their mode of binding differs from previously reported compounds, taking advantage of strong hydrophobic interaction. The new flexible biphenyl compounds show overall improved antiparasitic activity compared to rigid ones, with the best compound displaying a 2 nM antiplasmodial IC50 and suitable drug-like properties. This confirms the importance of compound flexibility for antimalarial activity and opens the way to new opportunities for antimalarial drug design.
Full text
Available for:
IJS, KILJ, NUK, UL, UM, UPUK
PDF
