Seeded Chain-Growth Polymerization of Proteins in Living Bacterial Cells.
Microbially produced protein-based materials (PBMs) are appealing due to use of renewable feedstock, low energy requirements, tunable side-chain chemistry, and biodegradability. However, high-strength PBMs typically have high molecular weights (HMW) and repetitive sequences that are difficult to microbially produce due to genetic instability and metabolic burden. We report the development of a biosynthetic strategy termed seeded chain-growth polymerization (SCP) for synthesis of HMW PBMs in living bacterial cells. SCP uses split intein (SI) chemistry to cotranslationally polymerize relatively small, genetically stable material protein subunits, effectively preventing intramolecular cyclization. We apply SCP to bioproduction of spider silk in Escherichia coli, generating HMW spider silk proteins (spidroins) up to 300 kDa, resulting in spidroin fibers of high strength, modulus, and toughness. SCP provides a modular strategy to synthesize HMW, repetitive material proteins, and may facilitate bioproduction of a variety of high-performance PBMs for broad applications.
['Bowen|Christopher Hyde|CH|', 'Reed|Tavis J|TJ|', 'Sargent|Cameron Jay|CJ|0000-0003-0442-9974', 'Mpamo|Beryl|B|', 'Galazka|Jonathan M|JM|', 'Zhang|Fuzhong|F|0000-0001-6979-7909']
D001704:Biopolymers D004926:Escherichia coli D005349:Fibroins D047668:Inteins D050296:Microbial Viability D008970:Molecular Weight D058105:Polymerization D017433:Protein Structure, Secondary D015203:Reproducibility of Results
chain-growth polymerization protein-based materials split inteins bioproduction spider silk synthetic biology