Authors:

Ethan A. Loredo

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About Paper:

Nanomaterials have vast potential in catalysis, drug delivery, and energy; however, it is difficult to engineer such materials with the necessary precision to implement at scale. Biotemplating offers a fine-tuned way to produce nanomaterials by allowing proteins to self-assemble into structures to be used for non-native purposes. The shell proteins PduA and PduJ from the Pdu (1,2-propanediol utilization) microcompartment (MCP) from Salmonella enterica Typhimurium LT2 are candidates for this purpose since they self-assemble into higher order structures when expressed in cell-free protein synthesis (CFPS). Though only different by 13 amino acids, PduA and PduJ form vastly different structures when expressed in vitro. Moreover, single amino acid substitutions vastly reshape morphology and size of these high order protein assemblies, creating structures ranging from tight packs of long fibrils to hexagonal disks coupled with ribbon clusters [1]. Previous work built Pdu materials by using single protein variants per assembly. Here, we aimed to fine-tune the morphology and size of these structures further through the incorporation of multiple Pdu subunits as building blocks. To that end, we expressed previously characterized PduJ-FLAG and PduA-FLAG mutants that form noticeably different structures (e.g. hexagons and fibrils) with each other to determine if their co-assembly led to a change in morphology compared to the individual components on their own. We found that co-assembly of PduA and PduJ mutants altered the size and morphology of the resulting structures. We also found that shell protein homologs from other organisms co-assemble with PduA and PduJ, opening doors for further fine-tuning assembly. Finally, we were able to engineer a mechanism to localize heterologous proteins to PduJ fibrils. These results pave the way for endowing new functionality to Pdu nanomaterials such as enzyme co-immobilization or biosensing in addition. to further elucidating design rules that govern Pdu assembly.

Source:

Northwestern University

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Co-authors:

Ethan A. Loredo