Proteins that bind to a target protein of interest, termed “binders,” are essential components of biological research reagents and therapeutics. Target proteins present multiple binding surfaces with ...

De novo binder discovery is unpredictable and inefficient due to a lack of quantitative understanding of protein-protein interaction (PPI) sequence-function landscapes. Here, we use our PANCS-Binder t...

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BindCraft, an open-source, automated pipeline for de novo protein binder design with experimental success rates of 10–100%, leverages AlphaFold2 weights to generate binders with nanomolar affinity wit...

Phage-assisted noncontinuous selection of protein binders (PANCS-Binders) allows multiple high-diversity protein libraries to each be screened against a panel of dozens of targets for high-throughput ...

Compartmentalization by organelles and the dynamic control of protein localization within these compartmentalized spaces are key mechanisms for regulating biological processes in eukaryotic cells. Here, we present a bottom-up approach for constructing cell-sized liposomes (giant unilamellar vesicles, GUVs) encapsulating an artificial organelle with chemically controlled protein localization. In this system, proteins fused to Escherichia coli dihydrofolate reductase are rapidly recruited on demand from the inner solution to the interior of a DNA-droplet-based (“nucleus”-like) organelle within GUVs upon addition of a synthetic, DNA-binding trimethoprim derivative to the external solution. By coupling this system with a sequence-specific protease, we constructed a synthetic cell platform that enables chemically induced, multistep cascade reactions─including protein relocalization, organelle-specific enzymatic activity, and product release from the organelle─that culminate in the control of synthetic-cell phenotypes, such as pore formation in the GUV membrane. This work provides a versatile platform for the bottom-up creation of eukaryotic-like synthetic cells with sophisticated and programmable functions.

Creating artificial organelles that sequester and release specific proteins in response to a small molecule in mammalian cells is an attractive approach for regulating protein function. In this work, ...