Design of a hyperstable 60-subunit protein icosahedron

Abstract

The icosahedron and the dodecahedron are the largest of the Platonic solids, and icosahedral protein structures are widely utilized in biological systems for packaging and transport1,2. There has been considerable interest in repurposing such structures3–5, for example, virus-like particles for the targeted delivery and vaccine design. The ability to design proteins that self assemble into precisely specified, highly ordered icosahedral structures would open the door to a new generation of protein 'containers' that could exhibit properties custom-made for various applications. In this manuscript, we describe the computational design of an icosahedral nano-cage that self-assembles from trimeric building blocks. Electron microscopy images of the designed protein expressed in E. coli reveals a homogenous population of icosahedral particles nearly identical to the design model. The particles are stable in 6.7 M guanidine hydrochloride at up to 80 °C, and undergo extremely abrupt, but reversible, disassembly between 2 M and 2.25 M guanidinium thiocyanate. The icosahedron is robust to genetic fusions: one or two copies of superfolder GFP can be fused to each of the 60 subunits to create highly fluorescent standard candles for light microscopy, and a designed protein pentamer can be placed in the center of each of the twenty pentameric faces to potentially gate macromolecule access to the nanocage interior. Such robust designed nanocages should have considerable utility for targeted drug delivery6, vaccine design7, and synthetic biology8.

Supercharging enables organized assembly of synthetic biomolecules

“…Control SuPrA protomers containing entirely Ceru+32, entirely GFP-17, or an octamer of Ceru+32 juxtaposed with an octamer of GFP-17 were unstable at all NaCl concentrations, consistent with the experimental observations ( Supplementary Fig. 12, 13) Finer-grained structural modeling (i.e., of hydrophobic effects and counterion release) must await specific structural and physical details that should become manifest with further imaging (i.e., cryo-EM) experiments that better determine the precise symmetric arrangement of monomers.While previous studies have shown that it is possible to design synthetic protein oligomers either by modifying extant structures [13][14][15][16] or de novo, [8][9][10][11][12] we now show that it may be possible to induce the formation of novel quaternary structures through the simple expedient of supercharging. In so doing, we demonstrate that precise molecular configurations can be obtained by focusing on the driving forces for assembly (charge:charge interactions) rather than the details of the interface.…”

“…While previous studies have shown that it is possible to design synthetic protein oligomers either by modifying extant structures [13][14][15][16] or de novo, [8][9][10][11][12] we now show that it may be possible to induce the formation of novel quaternary structures through the simple expedient of supercharging. In so doing, we demonstrate that precise molecular configurations can be obtained by focusing on the driving forces for assembly (charge:charge interactions) rather than the details of the interface.…”

“…[1][2][3][4][5][6][7] Repeated, symmetric interactions between molecular building blocks enable the formation of complex, defined, assemblies from a small total number of components as well as dynamic propagation of conformational change. [3][4][5][6][7][8] However, while de novo engineering of artificial symmetrical biomolecular complexes has begun to be explored, [9][10][11][12][13][14][15] these efforts generally rely upon precise design of molecular interfaces. 16,17 In contrast, studies of simple synthetic colloids suggests that higher order structures can be derived based solely on packing and energetic and considerations.…”

Nouvelles stratégies d’innovations vaccinales et leurs application en médecine vétérinaire

“…(Plummer & Manchester, 2011). Au-delà des capsides virales, de nombreuses autres protéines naturelles s'auto-assemblent spontanément, la ferritine par exemple étudiée dans des vaccins grippaux, la protéine E2 de Geobacillus stearothermophilus (López-Sagaseta et al 2016), des protéines de synthèse comme 13-01 aldolase (Hsia et al 2016), la lumazine synthase de la bactérie Aquifex aeolicus (Ra et al 2014). Le concept s'élargit de jour en jour avec des microsphères, liposomes, ou micelles naturelles ou synthétiques, des nano-émulsions, des nano-vésicules, des peptides supramoléculaires assemblés en nanofibres, etc… (Wen & Collier, 2015).…”

Novel Treatment Approach in Schizophrenia: Substitution of Glial Binding Proteins

“…The ability to design proteins that self-assemble into precisely specified, highly ordered icosahedral structures open the door to a new generation of proteins [19]. Recently, a new method for producing and isolating human glial progenitor cells has been developed enabling a cell-based therapy for neurological and neuropsychiatric disorders [20].…”