Abstract
Despite broad interest in understanding the biological implications of protein farnesylation in regulating different facets of cell biology, the use of this post-translational modification to develop protein-based materials and therapies remains underexplored. The progress has been slow due to the lack of accessible methodologies to generate farnesylated proteins with broad physicochemical diversities rapidly. This limitation, in turn, has hindered the empirical elucidation of farnesylated proteins’ sequence–structure–function rules. To address this gap, we genetically engineered prokaryotes to develop operationally simple, high-yield biosynthetic routes to produce farnesylated proteins and revealed determinants of their emergent material properties (nano-aggregation and phase-behavior) using scattering, calorimetry, and microscopy. These outcomes foster the development of farnesylated proteins as recombinant therapeutics or biomaterials with molecularly programmable assembly.
Highlights
The development of new protein-based biopharmaceuticals and materials is a vibrant area of research at the interface of chemistry, biology, and materials science and engineering.[1−8] Efforts in this space have traditionally focused on engineering the amino acid sequence and structure of proteins to achieve a specific function for applications including biomaterials,[9−12] sensors,[13] and biocatalysts,[14] among others.[15−17] Instead of changing the sequence of proteins, cells leverage an alternative strategy, post-translational modification (PTMs), to modulate a protein’s structure and function with exquisite spatiotemporal control.[18]
To reveal how farnesylation modulates the assembly and properties of proteins, we focused on elastin-like polypeptides (ELPs) as a model system
By converging synthetic biology with materials science, we developed recombinant platforms to produce farnesylated proteins with programmable assembly and temperaturedependent characteristics
Summary
The development of new protein-based biopharmaceuticals and materials is a vibrant area of research at the interface of chemistry, biology, and materials science and engineering.[1−8] Efforts in this space have traditionally focused on engineering the amino acid sequence and structure of proteins to achieve a specific function for applications including biomaterials,[9−12] sensors,[13] and biocatalysts,[14] among others.[15−17] Instead of changing the sequence of proteins, cells leverage an alternative strategy, post-translational modification (PTMs), to modulate a protein’s structure and function with exquisite spatiotemporal control.[18]. The unmodified (V/A)80-CVLL eluted at 9.8 min (dashed black curve), while (V/A)80-Fr eluted at 14.8 min (solid black curve) For both ELPs, the molecular ion peak corresponding to lipidated protein was shifted by m/z = +205.2 Da, consistent with adding a farnesyl motif (and removing a hydrogen atom from the thiol). We characterized the mesoscale assembly of farnesylmodified proteins above their Tt using differential interference contrast (DIC) microscopy Like their unmodified analogs, farnesylated proteins underwent liquid−liquid phase separation to form protein-rich coacervates (droplets), albeit farnesylation altered the average size (size-distribution) of coacervates (Figure S10)
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