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Self-assembly and modular functionalization of three-dimensional crystals from oppositely charged proteins.

Liljeström V, Mikkilä J, Kostiainen MA - Nat Commun (2014)

Bottom Line: Well-developed, especially DNA-based, methods for their preparation exist, yet most techniques concentrate on molecular and synthetic nanoparticle systems in non-biocompatible environment.Here we describe the self-assembly and characterization of binary solids that consist of crystalline arrays of native biomacromolecules.Importantly, the whole preparation process takes place at room temperature in a mild aqueous medium allowing the processing of delicate biological building blocks into ordered structures with lattice constants in the nanometre range.

View Article: PubMed Central - PubMed

Affiliation: 1] Biohybrid Materials Group, Department of Biotechnology and Chemical Technology, Aalto University, 00076 Aalto, Finland [2] Molecular Materials Group, Department of Applied Physics, Aalto University, 00076 Aalto, Finland.

ABSTRACT
Multicomponent crystals and nanoparticle superlattices are a powerful approach to integrate different materials into ordered nanostructures. Well-developed, especially DNA-based, methods for their preparation exist, yet most techniques concentrate on molecular and synthetic nanoparticle systems in non-biocompatible environment. Here we describe the self-assembly and characterization of binary solids that consist of crystalline arrays of native biomacromolecules. We electrostatically assembled cowpea chlorotic mottle virus particles and avidin proteins into heterogeneous crystals, where the virus particles adopt a non-close-packed body-centred cubic arrangement held together by avidin. Importantly, the whole preparation process takes place at room temperature in a mild aqueous medium allowing the processing of delicate biological building blocks into ordered structures with lattice constants in the nanometre range. Furthermore, the use of avidin-biotin interaction allows highly selective pre- or post-functionalization of the protein crystals in a modular way with different types of functional units, such as fluorescent dyes, enzymes and plasmonic nanoparticles.

No MeSH data available.


Related in: MedlinePlus

Aqueous electrostatic self-assembly of binary CCMV–avidin crystals.Agarose gel electrophoresis mobility shift assay (a), electrophoretic mobility (μe) and dynamic light scattering (b) show the formation of large assemblies, which can be disassembled or reassembled by increasing (to 50 mM NaCl) or decreasing the electrolyte concentration, respectively (c). a.u., arbitrary units. Count rate in decreasing cNaCl curve is multiplied by 0.72 for clarity. Volume-average size distribution measured from the free proteins and binary assemblies (d). Integrated SAXS curves (vertically offset for clarity) show the evolution of scattering curves at different electrolyte concentrations (e). The longest correlation length is achieved at 15 mM NaCl concentration, which is the optimum for crystal formation.
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f2: Aqueous electrostatic self-assembly of binary CCMV–avidin crystals.Agarose gel electrophoresis mobility shift assay (a), electrophoretic mobility (μe) and dynamic light scattering (b) show the formation of large assemblies, which can be disassembled or reassembled by increasing (to 50 mM NaCl) or decreasing the electrolyte concentration, respectively (c). a.u., arbitrary units. Count rate in decreasing cNaCl curve is multiplied by 0.72 for clarity. Volume-average size distribution measured from the free proteins and binary assemblies (d). Integrated SAXS curves (vertically offset for clarity) show the evolution of scattering curves at different electrolyte concentrations (e). The longest correlation length is achieved at 15 mM NaCl concentration, which is the optimum for crystal formation.

Mentions: Electrophoretic mobility, DLS and small-angle X-ray scattering (SAXS) experiments were first used to study the electrostatic binding between CCMV and avidin leading to the formation of large self-assembled protein crystals (Fig. 2). Gel electrophoresis mobility shift assay indicates that without avidin, the CCMV particles are free, negatively charged and consequently show a high electrophoretic mobility towards the cathode. When avidin/CCMV w/w ratio is increased, CCMV particles gradually lose their mobility, and at a ratio of 0.64 most of the virus particles are bound into larger complexes with very low electrophoretic mobility (Fig. 2a). The electrophoretic mobilities were further quantified with laser Doppler velocimetry. CCMV particles in a solution containing no or very small amounts of avidin were measured to have mobilities between −1.5 × 10−4 and −1.2 × 10−4 cm2 V−1 s−1. At an avidin/CCMV mass ratio between 0.16 and 0.32, the electrophoretic mobility shifts from negative to positive reaching a final value of 1.6 × 10−4 cm2 V−1 s−1 at an avidin/CCMV mass ratio of ~0.64 (Fig. 2b). Solvated ions screen the electric field of charged particles that makes the CCMV–avidin assemblies sensitive to variations in the ionic strength of the solution. DLS experiments showed that large CCMV–avidin complexes (Dh >1 μm) are formed at low ionic strength. However, they are rapidly disassembled when cNaCl is increased to ~50 mM (Fig. 2c,d). The disassembly of CCMV–avidin complexes due to increasing ionic strength is a reversible process, and the structures can reassemble when the ionic strength is decreased. The complex formation is rapid in all cases and takes place within minutes (Supplementary Fig. 1).


Self-assembly and modular functionalization of three-dimensional crystals from oppositely charged proteins.

Liljeström V, Mikkilä J, Kostiainen MA - Nat Commun (2014)

Aqueous electrostatic self-assembly of binary CCMV–avidin crystals.Agarose gel electrophoresis mobility shift assay (a), electrophoretic mobility (μe) and dynamic light scattering (b) show the formation of large assemblies, which can be disassembled or reassembled by increasing (to 50 mM NaCl) or decreasing the electrolyte concentration, respectively (c). a.u., arbitrary units. Count rate in decreasing cNaCl curve is multiplied by 0.72 for clarity. Volume-average size distribution measured from the free proteins and binary assemblies (d). Integrated SAXS curves (vertically offset for clarity) show the evolution of scattering curves at different electrolyte concentrations (e). The longest correlation length is achieved at 15 mM NaCl concentration, which is the optimum for crystal formation.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4109007&req=5

f2: Aqueous electrostatic self-assembly of binary CCMV–avidin crystals.Agarose gel electrophoresis mobility shift assay (a), electrophoretic mobility (μe) and dynamic light scattering (b) show the formation of large assemblies, which can be disassembled or reassembled by increasing (to 50 mM NaCl) or decreasing the electrolyte concentration, respectively (c). a.u., arbitrary units. Count rate in decreasing cNaCl curve is multiplied by 0.72 for clarity. Volume-average size distribution measured from the free proteins and binary assemblies (d). Integrated SAXS curves (vertically offset for clarity) show the evolution of scattering curves at different electrolyte concentrations (e). The longest correlation length is achieved at 15 mM NaCl concentration, which is the optimum for crystal formation.
Mentions: Electrophoretic mobility, DLS and small-angle X-ray scattering (SAXS) experiments were first used to study the electrostatic binding between CCMV and avidin leading to the formation of large self-assembled protein crystals (Fig. 2). Gel electrophoresis mobility shift assay indicates that without avidin, the CCMV particles are free, negatively charged and consequently show a high electrophoretic mobility towards the cathode. When avidin/CCMV w/w ratio is increased, CCMV particles gradually lose their mobility, and at a ratio of 0.64 most of the virus particles are bound into larger complexes with very low electrophoretic mobility (Fig. 2a). The electrophoretic mobilities were further quantified with laser Doppler velocimetry. CCMV particles in a solution containing no or very small amounts of avidin were measured to have mobilities between −1.5 × 10−4 and −1.2 × 10−4 cm2 V−1 s−1. At an avidin/CCMV mass ratio between 0.16 and 0.32, the electrophoretic mobility shifts from negative to positive reaching a final value of 1.6 × 10−4 cm2 V−1 s−1 at an avidin/CCMV mass ratio of ~0.64 (Fig. 2b). Solvated ions screen the electric field of charged particles that makes the CCMV–avidin assemblies sensitive to variations in the ionic strength of the solution. DLS experiments showed that large CCMV–avidin complexes (Dh >1 μm) are formed at low ionic strength. However, they are rapidly disassembled when cNaCl is increased to ~50 mM (Fig. 2c,d). The disassembly of CCMV–avidin complexes due to increasing ionic strength is a reversible process, and the structures can reassemble when the ionic strength is decreased. The complex formation is rapid in all cases and takes place within minutes (Supplementary Fig. 1).

Bottom Line: Well-developed, especially DNA-based, methods for their preparation exist, yet most techniques concentrate on molecular and synthetic nanoparticle systems in non-biocompatible environment.Here we describe the self-assembly and characterization of binary solids that consist of crystalline arrays of native biomacromolecules.Importantly, the whole preparation process takes place at room temperature in a mild aqueous medium allowing the processing of delicate biological building blocks into ordered structures with lattice constants in the nanometre range.

View Article: PubMed Central - PubMed

Affiliation: 1] Biohybrid Materials Group, Department of Biotechnology and Chemical Technology, Aalto University, 00076 Aalto, Finland [2] Molecular Materials Group, Department of Applied Physics, Aalto University, 00076 Aalto, Finland.

ABSTRACT
Multicomponent crystals and nanoparticle superlattices are a powerful approach to integrate different materials into ordered nanostructures. Well-developed, especially DNA-based, methods for their preparation exist, yet most techniques concentrate on molecular and synthetic nanoparticle systems in non-biocompatible environment. Here we describe the self-assembly and characterization of binary solids that consist of crystalline arrays of native biomacromolecules. We electrostatically assembled cowpea chlorotic mottle virus particles and avidin proteins into heterogeneous crystals, where the virus particles adopt a non-close-packed body-centred cubic arrangement held together by avidin. Importantly, the whole preparation process takes place at room temperature in a mild aqueous medium allowing the processing of delicate biological building blocks into ordered structures with lattice constants in the nanometre range. Furthermore, the use of avidin-biotin interaction allows highly selective pre- or post-functionalization of the protein crystals in a modular way with different types of functional units, such as fluorescent dyes, enzymes and plasmonic nanoparticles.

No MeSH data available.


Related in: MedlinePlus