Engineered shell proteins confer improved encapsulated pathway behavior in a bacterial microcompartment

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

doi:10.1101/106716

Cited by 3 publications

References 42 publications

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Programmed loading and rapid purification of engineered bacterial microcompartment shells

et al. 2018

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Bacterial microcompartments (BMCs) are selectively permeable proteinaceous organelles which encapsulate segments of metabolic pathways across bacterial phyla. They consist of an enzymatic core surrounded by a protein shell composed of multiple distinct proteins. Despite great potential in varied biotechnological applications, engineering efforts have been stymied by difficulties in their isolation and characterization and a dearth of robust methods for programming cores and shell permeability. We address these challenges by functionalizing shell proteins with affinity handles, enabling facile complementation-based affinity purification (CAP) and specific cargo docking sites for efficient encapsulation via covalent-linkage (EnCo). These shell functionalizations extend our knowledge of BMC architectural principles and enable the development of minimal shell systems of precisely defined structure and composition. The generalizability of CAP and EnCo will enable their application to functionally diverse microcompartment systems to facilitate both characterization of natural functions and the development of bespoke shells for selectively compartmentalizing proteins.

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Programmed loading and rapid purification of engineered bacterial microcompartment shells

et al. 2018

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A Catalog of the Diversity and Ubiquity of Metabolic Organelles in Bacteria

Sutter

Schulz

et al. 2021

Preprint

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Bacterial microcompartments (BMCs) are organelles that segregate segments of metabolic pathways that are incompatible with the surrounding metabolism. These metabolic modules consist entirely of protein, thus they are directly genetically encoded organelles. The BMC membrane is composed of families of proteins that oligomerize into pentagonal and hexagonal building blocks, typically perforated by pores, that tile into a polyhedral shell. The shell protein families are structurally homologous, with the function of the BMC determined by the encapsulated enzymes and the permeability properties of the constituent shell proteins. BMCs can be identified bioinformatically by locating genes encoding shell proteins, which are generally found proximal to those for the encapsulated enzymes. We here performed a large-scale sequence-based analysis of all shell proteins across the bacterial tree of life. With recent advances in genome-resolved metagenomics and the emphasis on “microbial dark matter”, many new genome sequences from diverse and obscure bacterial species clades have become available. We find that locus-specific designations of shell proteins should be supplanted, because higher level patterns of co-occurrence are evident. Moreover, the number of identifiable BMC loci has increased twenty-fold since the last comprehensive census of 2014. While we can assign many to bioinformatically characterized loci, the addition of new types uncovered in this study doubles the number of distinct BMC types described. In addition, we predict several new functional types that expand the range of catalysis encapsulated in BMCs, an intriguing example is an organelle for the degradation of an aromatic substrate, compartmentalized in an unusually simple shell, with potential for bioremediation. Our comprehensive survey of bacterial metabolic organelles underscores that there is compartmentalized dark biochemistry yet to be discovered through genome sequencing. The finding of up to six distinct BMC loci in a single genome underscores the role of BMCs in conferring metabolic flexibility and provides new insights into how certain clades have adapted to or even dominate, as in dysbiosis, an environmental niche. Our catalog provides a rich substrate for downstream experimental characterization of these metabolic modules and broadens the foundation for the development of BMC-based nanoarchitectures for biomedical and bioengineering applications.

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Enzymatically-active bacterial microcompartments follow substrate gradients and are protected from aggregation in a cell-free system

Steinkühler

Abrahamson

Agudo-Canalejo

et al. 2022

Preprint

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The ability to dynamically control organelle movement and position is essential for cellular function. Yet the underlying mechanisms driving this organization have not been fully resolved. Here, we draw from recent experimental observations and theoretical models of enzyme chemotaxis to demonstrate the chemotaxis of a bacterial organelle, the 1,2 propanediol (1,2-PD) utilization bacterial microcompartment (MCP) from Salmonella enterica. Upon encapsulating MCPs in a cell-like, biomimetic compartment, we observed the directed movement of MCPs along an external gradient of substrate. Our analysis shows that MCPs not only chemotax towards their substrate but also that enzymatic activity and substrate turnover protect them against large-scale aggregation. Our results provide a first experimental demonstration of organelle chemotaxis in a synthetic cellular system and support a recent theoretical model of chemotaxis. Together this work reveals a potentially significant driver of organelle organization while contributing to the construction of synthetic cell-like materials.

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Engineered shell proteins confer improved encapsulated pathway behavior in a bacterial microcompartment

Cited by 3 publications

References 42 publications

Programmed loading and rapid purification of engineered bacterial microcompartment shells

Programmed loading and rapid purification of engineered bacterial microcompartment shells

A Catalog of the Diversity and Ubiquity of Metabolic Organelles in Bacteria

Enzymatically-active bacterial microcompartments follow substrate gradients and are protected from aggregation in a cell-free system

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