Bottom-up Construction of a Primordial Carboxysome Mimic

Frey, Raphael; Mantri, Shiksha; Rocca, Marco; Hilvert, Donald

doi:10.1021/jacs.6b04744

Cited by 79 publications

(77 citation statements)

References 32 publications

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“…By providing a protective shell for molecular cargo, these nanostructures serve as multipurpose containers for the transport of viral payloads1, biomineralization2, protein folding3 and degradation4, enzyme encapsulation56 and even catalysis of short metabolic sequences78. Inspired by this precedent, protein cages are being engineered in the laboratory as customized nanoreactors910, delivery or display vehicles11 and artificial organelles1213 for diverse applications in medicine, materials science and synthetic biology.…”

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confidence: 99%

Structure and assembly of scalable porous protein cages

et al. 2017

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Proteins that self-assemble into regular shell-like polyhedra are useful, both in nature and in the laboratory, as molecular containers. Here we describe cryo-electron microscopy (EM) structures of two versatile encapsulation systems that exploit engineered electrostatic interactions for cargo loading. We show that increasing the number of negative charges on the lumenal surface of lumazine synthase, a protein that naturally assembles into a ∼1-MDa dodecahedron composed of 12 pentamers, induces stepwise expansion of the native protein shell, giving rise to thermostable ∼3-MDa and ∼6-MDa assemblies containing 180 and 360 subunits, respectively. Remarkably, these expanded particles assume unprecedented tetrahedrally and icosahedrally symmetric structures constructed entirely from pentameric units. Large keyhole-shaped pores in the shell, not present in the wild-type capsid, enable diffusion-limited encapsulation of complementarily charged guests. The structures of these supercharged assemblies demonstrate how programmed electrostatic effects can be effectively harnessed to tailor the architecture and properties of protein cages.

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confidence: 99%

Structure and assembly of scalable porous protein cages

et al. 2017

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“…In addition, modulating the chemical structure of nanostructure-forming monomers to preferentially exclude O 2 from these complexes may result in further enhancement of RubisCO’s Ω value. Likewise, co-immobilization with carbonic anhydrase might further enhance CO 2 availability around RubisCO molecules as in carboxysomes and analogous micro-compartments [44, 45]. …”

Section: Discussionmentioning

confidence: 99%

“…Cyanobacteria and some proteobacteria optimize CO 2 fixation by encapsulating RubisCO and carbonic anhydrase (CA) within intracellular compartments called carboxysomes; analogous micro-compartments are employed by eukaryotic algae [43, 44]. In a recent study, functional RubisCO was co-encapsulated with CA in synthetic carboxysome mimics, which could protect the enzyme from proteolytic degradation [45]. With these biological structures in mind, we postulated that RubisCO and other enzymes might be coaxed to self-assemble with nanostructures.…”

Section: Introductionmentioning

confidence: 99%

Synthetic CO2-fixation enzyme cascades immobilized on self-assembled nanostructures that enhance CO2/O2 selectivity of RubisCO

Satagopan

Sun

Parquette

et al. 2017

Biotechnol Biofuels

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BackgroundWith increasing concerns over global warming and depletion of fossil-fuel reserves, it is attractive to develop innovative strategies to assimilate CO2, a greenhouse gas, into usable organic carbon. Cell-free systems can be designed to operate as catalytic platforms with enzymes that offer exceptional selectivity and efficiency, without the need to support ancillary reactions of metabolic pathways operating in intact cells. Such systems are yet to be exploited for applications involving CO2 utilization and subsequent conversion to valuable products, including biofuels. The Calvin–Benson–Bassham (CBB) cycle and the enzyme ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) play a pivotal role in global CO2 fixation.ResultsWe hereby demonstrate the co-assembly of two RubisCO-associated multienzyme cascades with self-assembled synthetic amphiphilic peptide nanostructures. The immobilized enzyme cascades sequentially convert either ribose-5-phosphate (R-5-P) or glucose, a simpler substrate, to ribulose 1,5-bisphosphate (RuBP), the acceptor for incoming CO2 in the carboxylation reaction catalyzed by RubisCO. Protection from proteolytic degradation was observed in nanostructures associated with the small dimeric form of RubisCO and ancillary enzymes. Furthermore, nanostructures associated with a larger variant of RubisCO resulted in a significant enhancement of the enzyme’s selectivity towards CO2, without adversely affecting the catalytic activity.ConclusionsThe ability to assemble a cascade of enzymes for CO2 capture using self-assembling nanostructure scaffolds with functional enhancements show promise for potentially engineering entire pathways (with RubisCO or other CO2-fixing enzymes) to redirect carbon from industrial effluents into useful bioproducts.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-017-0861-6) contains supplementary material, which is available to authorized users.

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“…Cells commonly control metabolic pathways by encapsulating sequentially acting enzymes . The activities of ribulose‐1,5‐bisphosphate carboxylase/oxygenase and carbonic anhydrase enzyme were investigated upon coencapsulation in A. aeolicus LS‐13 cages . Tobacco etch virus protease with designed peptide substrates having different charged tags, including green fluorescent protein (+36), was encapsulated in A. aeolicus LS‐13 .…”

Section: Other Biomedical Applicationsmentioning

confidence: 99%

“…100,101 The activities of ribulose-1,5-bisphosphate carboxylase/oxygenase and carbonic anhydrase enzyme were investigated upon coencapsulation in A. aeolicus LS-13 cages. [102][103][104][105][106] Tobacco etch virus protease with designed peptide substrates having different charged tags, including green fluorescent protein (+36), was encapsulated in A. aeolicus LS-13. 102,107 The study elucidated cellular phenomena including ubiquitination, proteasomal degradation, and peptidic signaling sequences.…”

Section: Riboflavin-producing Microbiotamentioning

confidence: 99%

Understanding the riboflavin biosynthesis pathway for the development of antimicrobial agents

Kundu

Sarkar

Ray

et al. 2019

Medicinal Research Reviews

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Life on earth depends on the biosynthesis of riboflavin, which plays a vital role in biological electron transport processes. Higher mammals obtain riboflavin from dietary sources; however, various microorganisms, including Gram‐negative pathogenic bacteria and yeast, lack an efficient riboflavin‐uptake system and are dependent on endogenous riboflavin biosynthesis. Consequently, the inhibition of enzymes in the riboflavin biosynthesis pathway would allow selective toxicity to a pathogen and not the host. Thus, the riboflavin biosynthesis pathway is an attractive target for designing novel antimicrobial drugs, which are urgently needed to address the issue of multidrug resistance seen in various pathogens. The enzymes involved in riboflavin biosynthesis are lumazine synthase (LS) and riboflavin synthase (RS). Understanding the details of the mechanisms of the enzyme‐catalyzed reactions and the structural changes that occur in the enzyme active sites during catalysis can facilitate the design and synthesis of suitable analogs that can specifically inhibit the relevant enzymes and stop the generation of riboflavin in pathogenic bacteria. The present review is the first compilation of the work that has been carried out over the last 25 years focusing on the design of inhibitors of the biosynthesis of riboflavin based on an understanding of the mechanisms of LS and RS. This review aimed to address the fundamental advances in our understanding of riboflavin biosynthesis as applied to the rational design of a novel class of inhibitors. These advances have been aided by X‐ray structures of ligand‐enzyme complexes, rotational‐echo, double‐resonance nuclear magnetic resonance spectroscopy, high‐throughput screening, virtual screenings, and various mechanistic probes.

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Bottom-up Construction of a Primordial Carboxysome Mimic

Cited by 79 publications

References 32 publications

Structure and assembly of scalable porous protein cages

Structure and assembly of scalable porous protein cages

Synthetic CO2-fixation enzyme cascades immobilized on self-assembled nanostructures that enhance CO2/O2 selectivity of RubisCO

Understanding the riboflavin biosynthesis pathway for the development of antimicrobial agents

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