A5032403655- bioluminescence and chemiluminescence research
- Photoreceptor and optogenetics research
- Protease and Inhibitor Mechanisms
- Advanced biosensing and bioanalysis techniques
- Protein Structure and Dynamics
- Molecular Communication and Nanonetworks
- Advanced Biosensing Techniques and Applications
- Enzyme Production and Characterization
- Invertebrate Immune Response Mechanisms
- Glycosylation and Glycoproteins Research
- Photosynthetic Processes and Mechanisms
- Monoclonal and Polyclonal Antibodies Research
- Antiplatelet Therapy and Cardiovascular Diseases
- Enzyme Structure and Function
- Microbial Metabolism and Applications
- Amino Acid Enzymes and Metabolism
- Receptor Mechanisms and Signaling
- Pharmacological Effects of Natural Compounds
- Chemical Reactions and Isotopes
- Studies on Chitinases and Chitosanases
- Lipid Membrane Structure and Behavior
- Calpain Protease Function and Regulation
- Signaling Pathways in Disease
- Click Chemistry and Applications
- Protein purification and stability
VIB-UGent Center for Inflammation Research
2025
Masaryk University
2019-2024
Ghent University Hospital
2024
University Hospital Brno
2019-2023
St. Anne's University Hospital Brno
2019-2023
International Clinical Research Center, St. Anne's University Hospital Brno
2022
RECETOX
2019
Protein dynamics are often invoked in explanations of enzyme catalysis, but their design has proven elusive. Here we track the role evolution, starting from evolvable and thermostable ancestral protein AncHLD-RLuc which catalyses both dehalogenase luciferase reactions. Insertion-deletion (InDel) backbone mutagenesis challenged scaffold dynamics. Screening for activities reveals InDel mutations localized three distinct regions that lead to altered (based on crystallographic B-factors,...
To obtain structural insights into the emergence of biological functions from catalytically promiscuous enzymes, we reconstructed an ancestor distinct, but evolutionarily related, haloalkane dehalogenases (EC 3.8.1.5) and Renilla luciferase 1.13.12.5). This has both hydrolase monooxygenase activities. Its crystal structure solved to 1.39 Å resolution revealed presence a catalytic pentad conserved in dehalogenase descendants molecular oxygen bound between two residues typically stabilizing...
NanoLuc, a superior β-barrel fold luciferase, was engineered 10 years ago but the nature of its catalysis remains puzzling. Here experimental and computational techniques are combined, revealing that imidazopyrazinone luciferins bind to an intra-barrel catalytic site also allosteric shaped on enzyme surface. Structurally, binding prevents simultaneous site, vice versa, through concerted conformational changes. We demonstrate restructuration can boost luminescent reaction in remote active...
<title>Abstract</title> Despite recent advances in the computational design of protein binders, designing proteins that bind with high affinity to polar targets remains an outstanding problem. Here we show RFdiffusion can be conditioned efficiently generate scaffolds form geometrically matched extended beta-sheets target edge beta-strands which groups on are nearly perfectly complemented hydrogen bonding design. We use this approach binders against a set therapeutically relevant (KIT,...
Interleukin-36 receptor (IL-36R) is activated by IL-36α, IL-36β, and IL-36 γ to elicit pro-inflammatory signaling targeted in acute skin inflammation the approved antibody spesolimab. IL-37 was recently proposed as a new IL-36R agonist. Such diverse agonist repertoire together with antagonistic IL-36Ra IL-38 create fascinating structure-function landscape for IL-36R, albeit one that poorly understood. Here, we elucidate how grapples IL-36γ low affinity enable facile recruitment of shared...
Abstract Despite recent advances in the computational design of protein binders, designing proteins that bind with high affinity to polar targets remains an outstanding problem. Here we show RFdiffusion can be conditioned efficiently generate scaffolds form geometrically matched extended beta-sheets target edge beta-strands which groups on are nearly perfectly complemented hydrogen bonding design. We use this approach binders against a set therapeutically relevant (KIT, PDGFRɑ, ALK-2, ALK-3,...
The plasminogen activator staphylokinase is a fibrin-specific thrombolytic biomolecule and an attractive target for the development of effective myocardial infarction stroke therapy. To engineer protein rationally, detailed understanding biochemical mechanism limiting steps essential. Conventional fitting to equations derived on basis simplifying approximations may be inaccurate complex mechanisms such as that staphylokinase. We employed modern numerical approach global kinetic data analysis...
<p>Insertion-deletion mutations are sources of major functional innovations in naturally evolved proteins, but directed evolution methods rely primarily on substitutions. Here, we report a powerful strategy for engineering backbone dynamics based InDel mutagenesis stable and evolvable template, its validation application to thermostable ancestor haloalkane dehalogenase <i>Renilla</i> luciferase. First, extensive multidisciplinary analysis linked the conformational...
The engineering of efficient enzymes for large-scale production industrially relevant compounds is a challenging task. Utilizing rational protein design, which relies on comprehensive understanding mechanistic information, holds significant promise achieving success in this endeavor. Pre-steady-state kinetic measurements, obtained either through fast-mixing techniques or photoswitchable substrates, provide crucial insights. latter approach not only furnishes clarity but also affords...
Abstract The widely used coelenterazine-powered Renilla luciferase was discovered over 40 years ago but the oxidative mechanism by which it generates blue photons remains unclear. Here we decipher -type bioluminescence through crystallographic, spectroscopic, and computational experiments. Structures of ancestral extant luciferases complexed with substrate-like analogue azacoelenterazine or a reaction product were obtained, providing unprecedented snapshots coelenterazine-to-coelenteramide...
Cardio- and cerebrovascular diseases are leading causes of death disability, resulting in one the highest socio-economic burdens any disease type. The discovery bacterial human plasminogen activators their use as thrombolytic drugs have revolutionized treatment these pathologies. Fibrin-specific agents an advantage over non-specific factors because lower rates deleterious side effects. Specifically, staphylokinase (SAK) is a pharmacologically attractive indirect activator protein origin that...
Abstract NanoLuc, a superior β-barrel fold luciferase, was engineered 10 years ago but the nature of its catalysis remains puzzling. Here experimental and computational techniques were combined, revealing that imidazopyrazinone luciferins bind to an intra-barrel catalytic site also allosteric shaped on enzyme surface. Binding prevents simultaneous binding site, vice versa, through concerted conformational changes. We demonstrate restructuration can dramatically boost luminescent reaction in...
Insertion-deletion mutations are sources of major functional innovations in naturally evolved proteins, but directed evolution methods rely primarily on substitutions. Here, we report a powerful strategy for engineering backbone dynamics based InDel mutagenesis stable and evolvable template, its validation application to thermostable ancestor haloalkane dehalogenase Renilla luciferase. First, extensive multidisciplinary analysis linked the conformational flexibility loop-helix fragment...