<i>In Vivo</i> Encapsulation of Nucleic Acids Using an Engineered Nonviral Protein Capsid

Lilavivat, Seth; Sardar, Debosmita; Jana, Subrata; Thomas, Gary K.; Woycechowsky, Kenneth J.

doi:10.1021/ja302743g

Cited by 41 publications

(38 citation statements)

References 29 publications

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“…In addition, Hilvert and coworkers reported that engineered nonviral protein capsids such as lumazine synthase from Aquifex aeolicus are also applicable as nano-carriers for proteins and DNA. [14][15][16] Currently, however, functional modification of natural protein capsules, such as plant viral capsids and virus-like lumazine synthase capsids, requires complicated technologies such as gene recombination and protein expression. Therefore, the reconstruction of virus-like nanocapsules from synthetic molecules would enhance their potential for molecular designs.…”

Section: Introductionmentioning

confidence: 99%

Guest-binding behavior of peptide nanocapsules self-assembled from viral peptide fragments

Matsuura

Watanabe

Matsushita

et al. 2013

Polym J

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The binding behavior of guests (dyes and DNA) into peptide nanocapsules formed via self-assembly of a 24-mer b-annulus peptide fragment obtained from the capsid protein of the tomato bushy stunt virus is reported. The pH dependence of the f potential of the peptide nanocapsules indicates that the C-and N-termini are directed to the exterior and interior of the nanocapsules, respectively. Equilibrium dialysis experiments with dyes and the peptide nanocapsules at pH 7 showed that the peptide nanocapsules tend to bind anionic dyes. Binding of sodium 8-anilinonaphthalene-1-sulfonate and uranine into the peptide nanocapsules minimally affected the size of the nanocapsules, whereas binding of other anionic dyes resulted in the formation of precipitates. In addition, binding of Thioflavin T to the b-annulus peptide promoted disassembly of the nanocapsules. Complexation of the b-annulus peptide with M13 phage DNA formed a core-shell nanosphere in which the DNA was encapsulated in the peptide assembly.

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Section: Introductionmentioning

confidence: 99%

Guest-binding behavior of peptide nanocapsules self-assembled from viral peptide fragments

Matsuura

Watanabe

Matsushita

et al. 2013

Polym J

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“…In the nearly 40 years since the first high-resolution structure of an icosahedral virus 11 , the structures and functions of a wide array of viral capsids have been characterized. This has inspired efforts to reengineer naturally occurring protein containers 12 and to design new polypeptides 13 to package biological molecules. In one case, lumazine synthase—a naturally occurring, non-viral protein container—was evolved in E. coli to sequester a toxic protein 14 .…”

mentioning

confidence: 99%

Evolution of a designed protein assembly encapsulating its own RNA genome

Butterfield

Lajoie

Gustafson

et al. 2017

Nature

185

237

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The challenges of evolution in a complex biochemical environment—coupling genotype to phenotype and protecting the genetic material—are solved elegantly in biological systems by nucleic acid encapsulation. In the simplest examples, viruses use capsids to surround their genomes. While these naturally occurring systems have been modified to change their tropism1 and to display proteins or peptides2–4, billions of years of evolution have favored efficiency at the expense of modularity, making viral capsids difficult to engineer. Synthetic systems composed of non-viral proteins could provide a “blank slate” to evolve desired properties for drug delivery and other biomedical applications, while avoiding the safety risks and engineering challenges associated with viruses. Here we create synthetic nucleocapsids—computationally designed icosahedral protein assemblies5, 6 with positively charged inner surfaces capable of packaging their own full-length mRNA genomes—and explore their ability to evolve virus-like properties by generating diversified populations using Escherichia coli as an expression host. Several generations of evolution resulted in drastically improved genome packaging (>133-fold), stability in whole murine blood (from less than 3.7% to 71% of packaged RNA protected after 6 hours of treatment), and in vivo circulation time (from less than 5 minutes to 4.5 hours). The resulting synthetic nucleocapsids package one full-length RNA genome for every 11 icosahedral assemblies, similar to the best recombinant adeno-associated virus (AAV) vectors7, 8. Our results show that there are simple evolutionary paths through which protein assemblies can acquire virus-like genome packaging and protection. Considerable effort has been directed at “top-down” modification of viruses to be safe and effective for drug delivery and vaccine applications1, 9, 10; the ability to computationally design synthetic nanomaterials and to optimize them through evolution now enables a complementary “bottom-up” approach with considerable advantages in programmability and control.

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“…Inspired by this, various classes of viral‐mimicking biohybrid NPs have been introduced for genome delivery . Understanding the structure of viruses inspired many efforts to engineer protein containers and to synthesize nucleic acid‐encapsulating polypeptides . It has recently been shown that protein assemblies could be re‐engineered to encapsulate their own genome .…”

Section: Challenges and Limitations For In Vivo Delivery Of Npsmentioning

confidence: 99%

Biohybrid Nanoparticles to Negotiate with Biological Barriers

Mohammadi

Corbo

Molinaro

et al. 2019

Small

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Incapability of effective cross-talk with biological environments has partly impaired the in vivo functionality of nanoparticles (NPs). Homing, biodistribution, and function of NPs could be engineered through regulating their interactions with in vivo niches. Inspired by communications in biological systems, endowing a "biological identity" to synthetic NPs is one approach to control their biodistribution, and immunonegotiation profiles. This syntheticbiological combination is referred to as biohybrid NPs, which comprise both i) engineerable, readily producible, and trackable synthetic NPs as well as ii) biological moieties with the capability to cross-talk with immunological barriers. Here, the latest understanding on the in vivo interactions of NPs, biological barriers they face, and emerging methods for quantitative measurements of NPs' biodistribution are reviewed. Some key biomolecules that have emerged as negotiators with the immune system in the context of cancer and autoimmunity, and their inspirations on biohybrid NPs are introduced. Critical design considerations for efficient cross-talk between NPs and innate and adaptive immunity followed by hybridization methods are also discussed. Finally, clinical translation challenges and future perspectives regarding biohybrid NPs are discussed.

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In Vivo Encapsulation of Nucleic Acids Using an Engineered Nonviral Protein Capsid

Cited by 41 publications

References 29 publications

Guest-binding behavior of peptide nanocapsules self-assembled from viral peptide fragments

Guest-binding behavior of peptide nanocapsules self-assembled from viral peptide fragments

Evolution of a designed protein assembly encapsulating its own RNA genome

Biohybrid Nanoparticles to Negotiate with Biological Barriers

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