Biological Containers: Protein Cages as Multifunctional Nanoplatforms

Uchida, Masaki; Klem, Michael T.; Allen, Mark; Suci, Peter A.; Flenniken, Michelle L.; Gillitzer, Eric; Varpness, Zachary; Liepold, Lars; Young, Mark; Douglas, Trevor

doi:10.1002/adma.200601168

Cited by 530 publications

(524 citation statements)

References 187 publications

(203 reference statements)

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“…1,2 Moreover, protein-protein interactions are also integral to the generation of cellular selfassembled nano-structures, and establishing how they control self-assembly could lead not only to fundamental understanding but also to the eventual rational design of novel structures for a myriad of applications such as the templation of inorganic nano-materials and for encapsulated reaction chemistry. [3][4][5][6][7][8] Although protein-protein interactions are intriguing medicinal targets, they have only recently been pursued for drug development studies somewhat owing to the discovery that although they often involve large, buried surface area, they can be inhibited using low molecular weight small molecules that target ''hot spot'' residues where the binding energy is concentrated. 9,10 Alanine shaving, where individual side changes are conceptually shaved to a methyl residuum and where the stabilities of the resulting mutants are determined with respect to wild type, is the most common method to identify hot spot residues.…”

Section: Introductionmentioning

confidence: 99%

Rational disruption of the oligomerization of the mini‐ferritin E. coli DPS through protein‐protein interface mutation

Zhang

Chee

et al. 2011

Protein Science

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DNA-binding protein from starved cells (DPS), a mini-ferritin capable of self-assembling into a 12-meric nano-cage, was chosen as the basis for an alanine-shaving mutagenesis study to investigate the importance of key amino acid residues, located at symmetry-related protein-protein interfaces, in controlling protein stability and self-assembly. Nine mutants were designed through simple inspection, synthesized, and subjected to transmission electron microscopy, circular dichroism, size exclusion chromatography, and ''virtual alanine scanning'' computational analysis. The data indicate that many of these residues may be hot spot residues. Most remarkably, two residues, R83 and R133, were observed to shift the oligomerization state to~50% dimer. Based on the hypothesis that these two residues constitute a ''hot strip,'' located at the ferritin-like threefold axis, the double mutant was generated which completely shuts down detectable formation of 12-mer in solution, favoring a cooperatively folded dimer. The fact that this effect logically builds upon the single mutants emphasizes that complex self-assembly has the potential to be manipulated rationally. This study should have an impact on the fundamental understanding of the assembly of DPS protein cages specifically and protein quaternary structure in general. In addition, as there is much interest in applying these and similar systems to the templation of nano-materials and drug delivery, the ability to control this ferritin's oligomerization state and stability could prove especially valuable.

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Section: Introductionmentioning

confidence: 99%

Rational disruption of the oligomerization of the mini‐ferritin E. coli DPS through protein‐protein interface mutation

Zhang

Chee

et al. 2011

Protein Science

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“…Such viruses can also be thought of as tunable colloids or nanoscale scaffolds. 14,15 Control over placement of…”

Section: -4 45mentioning

confidence: 99%

Incorporating stimulus-responsive character into filamentous virus assemblies

Bermudez

Hathorne

2008

Faraday Discuss.

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[DO NOT ALTER/DELETE THIS TEXT] 5Controlling interactions between building blocks, in either guided or selfassemblies, is becoming increasingly important for creation of functional materials. We have focused our attention on the well-known model assembly, the filamentous bacteriophage, where our strategy is to selectively alter surface features by focusing on spatially distinct capsid 10 proteins. Towards introducing stimulus-responsive behavior in these flexible, rod-like particles, we have introduced elastin-like polypeptide (ELP) motifs of isoleucine and tyrosine "guest" residues by recombinant DNA methods. Our hypothesis is that modification of the major coat capsid protein would be greatly amplified by the 2700 copies per particle. 15Characterization of ELP-phage particles was carried out by microbiological assays, zeta potential, dynamic light scattering, and calorimetry. Bacteria producing ELP-phage particles grow more slowly and surprisingly, ELPmodified phages display a significant reduction in viral infectivity. For the lengths of ELP inserts studied, modified phages do not aggregate from 20 solution as monitored by DLS. However, the hydrodynamic size of the phages depends on the details of the ELP motif.Zeta potential measurements reveal the particles are electrostatically stabilized, and this contributes in part to the energetic barrier against aggregation. Preliminary calorimetric data indicate subtle thermal transitions in the range 35-40 o C, 25 suggesting that the ELP motif may collapse without triggering macroscopic aggregation. The results are consistent with the classical picture of critical solution phenomena at low concentrations, where to drive phase separation, solvent quality must be increasingly poor. Apart from being model systems to study basic questions of self-assembly, extending the these modular 30 systems is likely to result in improved understanding and control over selfassembly in various applications.

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“…[6][7][8][9][10][11][12][13] However, it is still challenging to manipulate their self-assembly in a controlled way and to analyze their assembled products precisely at the molecular level. 14,15 In this study, we have generated two different individual mutants of a protein cage with functional groups either inside or outside of the cage ( Figure 1A).…”

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confidence: 99%

Controlled Assembly of Bifunctional Chimeric Protein Cages and Composition Analysis Using Noncovalent Mass Spectrometry

et al. 2008

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Protein cage architectures, such as viral capsids, ferritins, and heat shock proteins (Hsp), have been extensively used as model systems to study the self-assembly of macromolecular complexes [1][2][3][4][5] and as nanoreactors for materials synthesis. [6][7][8][9][10][11][12][13] However, it is still challenging to manipulate their self-assembly in a controlled way and to analyze their assembled products precisely at the molecular level. 14,15 In this study, we have generated two different individual mutants of a protein cage with functional groups either inside or outside of the cage ( Figure 1A). We chemically modified different cages and reconstructed chimeric cages with a controlled ratio of two subunit types ( Figure 1B). Using mass spectrometry, we were able to determine the compositions of the ensemble population and also of the individual chimeric cages within the population at the molecular level. A model based on a binomial distribution suggested chimeric cages are assembled by random incorporation of the two individual subunits.Mass spectrometry has been used to monitor multicomponent systems, because it can simultaneously resolve individual molecular masses present in a mixture. [16][17][18] Using a combination of electrospray ionization (ESI) 19 and a time-of-flight (TOF) mass analyzer, it is possible to determine the masses of individual protein components of a noncovalently associated macromolecular complex as well as the mass of the intact macromolecular complex without disturbing the structures. 16,[20][21][22][23] The Dps (DNA binding protein from starved cells) from the Gram-positive bacterium Listeria innocua (Li) is a member of the ferritin superfamily and prevents oxidative damage of DNA by accumulating iron atoms within its central cavity to produce an iron oxide core similar to that of ferritins. 24,25 The LiDps consists of 12 identical 18 kDa subunits that self-assemble into a hollow protein cage having tetrahedral 23 symmetry ( Figure 1A). 24 The LiDps has an outer diameter of 9 nm and an inner cavity diameter of 5 nm with 0.8 nm pores at the 3-fold axis where molecules can pass through to the interior ( Figure 1A). 24 The LiDps has been used as a template for nanomaterials synthesis of metal oxides of iron 26 and cobalt 27 as well as cadmium sulfide 28 and platinum 29 with or without modifications. The small number of subunits, robustness at high temperature, 26,27 and intrinsic biomineralizing capability 25 of the LiDps protein cage make it an attractive modifiable nanoreactor for nanomaterials syntheses. In addition, the defined small cavity size 24 allows synthesis of extremely small nanostructured materials. 29 Two individual cysteine mutants, one exposed on the interior surface and the other on the exterior surface, were generated to adapt the LiDps for selective chemical modifications. The serine residue at position 138 located in the middle of helix E where it is directed toward the inside cavity was substituted with cysteine (S138C) 29 ( Figure 1A, blue). Alternatively, for th...

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Biological Containers: Protein Cages as Multifunctional Nanoplatforms

Cited by 530 publications

References 187 publications

Rational disruption of the oligomerization of the mini‐ferritin E. coli DPS through protein‐protein interface mutation

Rational disruption of the oligomerization of the mini‐ferritin E. coli DPS through protein‐protein interface mutation

Incorporating stimulus-responsive character into filamentous virus assemblies

Controlled Assembly of Bifunctional Chimeric Protein Cages and Composition Analysis Using Noncovalent Mass Spectrometry

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