What if protein function could be triggered only when the right cell cues are present? With SMART, we combine split inteins and logic gating to create a programmable system for cell-specific protein ligation, thus opening new doors for synthetic biology and cell targeting.
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SMART relies on the fundamental AND-gated nature of split-inteins. When caged appropriately, we can use epitopes on the cell surface to drive the ligation of two inactive protein fragments to reconstitute a functional protein.
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This project started with the development of a split-SpyCatcher that is faithfully reconstituted when two distinct epitopes on cell surfaces is targeted.
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We are also able to develop other logic gated operations using this system. By engineering split-intein cages and SpyCatcher, we show that our system is able to operate a 3-input AND/NOT gate.
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However, given the widespread applications of AND-gated logic, we chose to focus on efforts on this front, and showed that we can do targeted cell depletion with exquisite selectivity for cells displaying the right combination of epitopes
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We are also able to show that SMART works with a range of targeting vectors, ranging from scFvs to peptides to small molecules. We also show that we can perform AND-gated enzymatic proximity labeling using APEX2 and photocatalytic proximity labeling using micro-Map
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While our prior applications of SMART involve a splice-and-retain mechanism, we also show that we can engineer SMART to perform a novel AND-gated splice-and-release operation. This is applied to the reconstitution of an active interleukin 1B from inactive fragments.
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We anticipate that this technology will enable targeted formation of biomolecules with reduced off-target toxicity. This paper is the start of many applications for SMART and stay tuned for more!
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Congratulations to Christian, Girum, Nick, Xuanjia, and Yutong! Link: https://t.co/KoOKDGH6Pd
@PrincetonChem @PrincetonInnov
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@MuirLab The inclusion of peptides as targeting vectors is particularly interesting. Are there specific considerations for their stability or delivery in this context?
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