<i>De novo</i> design of signal sequences to localize cargo to the 1,2‐propanediol utilization microcompartment

Jakobson, Christopher M.; Lee, Marilyn F. Slininger; Tullman‐Ercek, Danielle

doi:10.1002/pro.3144

Cited by 34 publications

(32 citation statements)

References 31 publications

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“…There is typically only one or two encapsulation peptides associated with each BMC type, and they may compete for the same shell protein binding partner 130 . The low encapsulation efficiency of non-native cargo that has been observed in several studies 40, 63,67,134,135 may also be due to the absence of additional protein domain interactions that contribute to the robustness of encapsulation. Several strategies have been explored to overcome these limitations.…”

Section: Bacterial Microcompartments In Bioengineeringmentioning

confidence: 95%

“…For example, encapsulation efficiency can be improved by delayed expression of the cargo relative to the shell proteins 136 and by altering the primary structure of encapsulation peptides 137 . To identify the primary structure features that are important for encapsulation efficiency, encapsulation peptides were designed de novo 134 and reporter proteins that were fused to encapsulation peptides from GRM and EUT BMCs were encapsulated in a PDU BMC 138 , demonstrating the potential for using encapsulation peptides to attach proteins to diverse BMC shells.…”

Section: Bacterial Microcompartments In Bioengineeringmentioning

confidence: 99%

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Bacterial microcompartments

et al. 2018

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Bacterial microcompartments (BMCs) are self-assembling organelles that consist of an enzymatic core that is encapsulated by a selectively permeable protein shell. The potential to form BMCs is widespread, found across the Kingdom Bacteria. BMCs have crucial roles in carbon dioxide fixation in autotrophs and the catabolism of organic substrates in heterotrophs. They contribute to the metabolic versatility of bacteria, providing a competitive advantage in specific environmental niches. Although BMCs were first visualized more than sixty years ago, it is mainly in the last decade that progress has been made in understanding their metabolic diversity and the structural basis of their assembly and function. This progress has not only heightened our understanding of their role in microbial metabolism but it is also beginning to enable their use in a variety of applications in synthetic biology. In this Review, we focus on recent insights into the structure, assembly, diversity and function of BMCs.

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Section: Bacterial Microcompartments In Bioengineeringmentioning

confidence: 95%

Section: Bacterial Microcompartments In Bioengineeringmentioning

confidence: 99%

Bacterial microcompartments

et al. 2018

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“…Furthermore, in addition to protein scaffolds, DNA scaffolds [ 12 , 13 ] and lipid-containing scaffolds [ 14 ] are also applied to improve the pathway efficiency. In addition to scaffolds, enzymes were also located to bacterial microcompartments (MCPs), such as ethanolamine utilization (Eut) [ 15 ] and 1,2-propanediol utilization (Pdu) MCPs for similar purposes [ 16 ]. The key point of these strategies was to localize the enzymes as close as possible to their substrates.…”

Section: Introductionmentioning

confidence: 99%

Optimizing the localization of astaxanthin enzymes for improved productivity

Zhu

et al. 2018

Biotechnol Biofuels

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BackgroundOne important metabolic engineering strategy is to localize the enzymes close to their substrates for improved catalytic efficiency. However, localization configurations become more complex the greater the number of enzymes and substrates is involved. Indeed, optimizing synthetic pathways by localizing multiple enzymes remains a challenge. Terpenes are one of the most valuable and abundant natural product groups. Phytoene, lycopene and β-carotene serve as common intermediates for the synthesis of many carotenoids and derivative compounds, which are hydrophobic long-chain terpenoids, insoluble in water and usually accumulate in membrane compartments.ResultsWhile β-ionone synthesis by β-carotene cleavage dioxygenase PhCCD1 and astaxanthin synthesis by β-carotene ketolase (CrtW) and β-carotene hydroxylase (CrtZ) differ in complexity (single and multiple step pathways), the productivity of both pathways benefited from controlling enzyme localization to the E. coli cell membrane via a GlpF protein fusion. Especially, the astaxanthin synthesis pathway comprises both CrtW and CrtZ, which perform four interchangeable reactions initiated from β-carotene. Up to four localization strategies of CrtW and CrtZ were exhaustively discussed in this work, and the optimal positioning strategy was achieved. CrtW and CrtZ were linked using a flexible linker and localized to the membrane via a GlpF protein fusion. Enzymes in the optimal localization configuration allowed a 215.4% astaxanthin production increase.ConclusionsThis work exploits a localization situation involving membrane-bound substrates, intermediates and multiple enzymes for the first time, and provides a workable positioning strategy to solve problems in similar circumstances.Electronic supplementary materialThe online version of this article (10.1186/s13068-018-1270-1) contains supplementary material, which is available to authorized users.

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“…Although these scaffolds have been manufactured in numerous recombinant expression systems (e.g., various prokaryotic and eukaryotic organisms), they have been mainly assembled in Escherichia coli strains (Diaz et al, 2018). Advances in protein engineering in recent years harnessed the unique structural assembly of EZPs to enable the de novo and in silico design of novel EZPs (King et al, 2014;Bale et al, 2016;Hsia et al, 2016;Jakobson et al, 2017;Xu et al, 2019).…”

Section: Enzyme-derived Nanoparticles (Ezps)mentioning

confidence: 99%

Bioengineered Polyhydroxyalkanoates as Immobilized Enzyme Scaffolds for Industrial Applications

Wong

Ogura

Chen

et al. 2020

Front. Bioeng. Biotechnol.

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Enzymes function as biocatalysts and are extensively exploited in industrial applications. Immobilization of enzymes using support materials has been shown to improve enzyme properties, including stability and functionality in extreme conditions and recyclability in biocatalytic processing. This review focuses on the recent advances utilizing the design space of in vivo self-assembled polyhydroxyalkanoate (PHA) particles as biocatalyst immobilization scaffolds. Self-assembly of biologically active enzyme-coated PHA particles is a one-step in vivo production process, which avoids the costly and laborious in vitro chemical cross-linking of purified enzymes to separately produced support materials. The homogeneous orientation of enzymes densely coating PHA particles enhances the accessibility of catalytic sites, improving enzyme function. The PHA particle technology has been developed into a remarkable scaffolding platform for the design of cost-effective designer biocatalysts amenable toward robust industrial bioprocessing. In this review, the PHA particle technology will be compared to other biological supramolecular assembly-based technologies suitable for in vivo enzyme immobilization. Recent progress in the fabrication of biological particulate scaffolds using enzymes of industrial interest will be summarized. Additionally, we outline innovative approaches to overcome limitations of in vivo assembled PHA particles to enable finetuned immobilization of multiple enzymes to enhance performance in multi-step cascade reactions, such as those used in continuous flow bioprocessing.

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De novo design of signal sequences to localize cargo to the 1,2‐propanediol utilization microcompartment

Cited by 34 publications

References 31 publications

Bacterial microcompartments

Bacterial microcompartments

Optimizing the localization of astaxanthin enzymes for improved productivity

Bioengineered Polyhydroxyalkanoates as Immobilized Enzyme Scaffolds for Industrial Applications

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