Selfassembling protein nanostructures as the new platform for
Self-assembling protein nanostructures as the new platform for designed biomimetic smart materials Dr. Helena GRADIŠAR NIC, Department of synthetic biology and immunology EN-FIST, Centre of excellence CENTRE OF EXCELLENCE
Department of synthetic biology and immunology
Department of synthetic biology and immunology Synthetic biology Nature Chemical Biology Nature Communications - Designed protein nanostructures Nucleic Acid Research Molecular Therapy - DNA-based biosynthetic scaffolds - Design of cellular logic circuits in mammalian cells - Medical applications of synthetic biology Nature Structural and Molecular Biology Nature Medicine Blood Molecular mechanism of innate immunity - Mechanism of signaling of Toll-like receptors - Mechanism of activation of cytosolic inflammasomes - My. D 88 in cancer and autoimmunity - Translational medicine: inhibitors, activators for vaccines and cancer therapy
Protein nanostructures – rapid development Luo et al. , Protein assembly: versatile approaches to construct highly ordered nanostructures, Chem Reviews, 2016
Modular topological nano-cage – INNOVATIVE STRATEGY Luo et al. , Protein assembly: versatile approaches to construct highly ordered nanostructures, Chem Reviews, 2016
Programmable building blocks – Coiled-coils Parallel dimer Antiparallel dimer We establish a set of orthogonal coiled coilforming peptide pairs.
Design of tetrahedron- like cage from a single chain Sequential order of modules Deconstruction of a polyhedron into rigid modules Toolbox of designed coil-forming modules Advantages: - precise positioning of functional groups - cavity bounded by coiled-coils - adjustable shape and size Selfassembling nanostructure
Production and isolation of polypeptide Amino acid sequence MKQLEKELQAIEKQLAQLQWKAQARKKKLAQLKKKLQA SGPGSPEDEIQQLEEEIAQLEQKNAALKEKNQALK YGSGPGDIEQELERAKASIRRLEQEVNQERSRMAYLQTLLAKSGPGQLEDKVEELLSKNYHLENEVARLKKLVGSGPGMKQ LEKELKQLEKELQAIEKQLAQLQWKAQARKKKLAQLKKKLQASGPGSPEDEIQALEEKNAQLKQEIAALEEKNQALKYGS GPGQLEDKVEELLSKNYHLENEVARLKKLVGSGPGSPEDKIAQLKQKIQALKQENQQLEEENAALEYG SGPGSPEDENAAL EEKIAQLKQKNAALKEEIQALEYGSGPGSPEDKIAQLKEENQQLEQKIQALKEENAALEYGSGPGDIEQELERAKASIRRLEQ EVNQERSRMAYLQTLLAKSGPGSPEDKNAALKEEIQALEEENQALEEKIAQLKYGHHHH Production in E. coli Isolation Purity
Characterization of structure Helical content Chemical stability Size determination Confirmation of tetrahedral nanostructure Gradišar et al. , Nature Chemical Biology, 2013 Comments in Nature, Science, Nature Biotechnology, C&E News… Visuatization by TEM and AFM
Coiled-Coil Protein Origami Design platform (Co. POD) https: //github. com/NIC-SBI/protein_origami
Next generation of self-assembling polyhedral cages
In vivo folding of nanocage – advantage for applications BMDM High biocompatibility – biomedical applications
Functionalization of designed polyhedra smart materials tethered assemblies shape and size of the scaffold and cavity regulated assembly/disassembly engineered binding and functional sites outside and/or inside of the cavity
Smart materials medical and biotechnological applications - Targeted delivery to selected tissue - Designed vaccines - Enzyme encapsulation - Biocatalysis - Smart functional materials - Sensors - Switches -. .
Conclusions - Polyhedral protein nanostructures – advanced biomaterials with a great potential for apllications - Production of is cost-effective and sustainable - Current challenges are the size and functionalization of designed nanostructures - Great expertise and infrastructure at NIC
Projects SRA (ARRS) projects ERA Syn. Bio (2014 -2017) Bio. Molecular Origami: establishing foundations in structural synthetic biology to engineer biomolcules for new routes to nanoscale objects and biomaterials Coordinator: prof. Jerala NIC; Partners: Oxford, Bristol, TUM, UW Seattle, UCLA M-Era. NET (2016 -2019) Medi. SURF: Designed nanostructured bioactive surfaces for precision medicines Partners: Leiden, Eindhoven, Bilbao, NIC
Acknowledgement Prof. Dr. Roman JERALA, Head of Department Dr. Ajasja Ljubetič Fabio Lapenta Dr. Igor Drobnak Jana Aupič Žiga Strmšek Dr. Andreja Majerle Nuša Krivec Mammalian cells: Dr. Mojca Benčina Dr. Iva Hafner Bratkovič Mice: Duško Lainšček Ex-colleagues: Dr. Vid Kočar Dr. Sabina Božič Tibor Doles Karen Butina Prof. Dr. Tomaž Pisanski Dr. Nino Bašič
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