Abstract
Manmade high-performance polymers are typically non-biodegradable and derived from petroleum feedstock through energy intensive processes involving toxic solvents and byproducts. While engineered microbes have been used for renewable production of many small molecules, direct microbial synthesis of high-performance polymeric materials remains a major challenge. Here we engineer microbial production of megadalton muscle titin polymers yielding high-performance fibers that not only recapture highly desirable properties of natural titin (i.e., high damping capacity and mechanical recovery) but also exhibit high strength, toughness, and damping energy — outperforming many synthetic and natural polymers. Structural analyses and molecular modeling suggest these properties derive from unique inter-chain crystallization of folded immunoglobulin-like domains that resists inter-chain slippage while permitting intra-chain unfolding. These fibers have potential applications in areas from biomedicine to textiles, and the developed approach, coupled with the structure-function insights, promises to accelerate further innovation in microbial production of high-performance materials.
Highlights
Manmade high-performance polymers are typically non-biodegradable and derived from petroleum feedstock through energy intensive processes involving toxic solvents and byproducts
We hypothesized that expressing this chimeric protein in E. coli would, through multiple rounds of intracellular SI-catalyzed ligation of 4Ig subunits, produce ultra-high molecular weight (UHMW) titin polymers (Fig. 1b)
Scanning transmission electron microscopy (STEM) of the purified titin polymer (Fig. 2b, Supplementary Fig. 3) showed the presence of numerous nanoscale fibrils with apparent chain-of-beads structures and cross-sectional diameters (6.1 ± 1.2 nm, Supplementary Fig. 4) that were similar to those previously observed for natural titin proteins[18]. These results suggest that our microbial production system can synthesize UHMW titin polymers with a substantial degree of folded structures similar to natural Ig domains
Summary
Manmade high-performance polymers are typically non-biodegradable and derived from petroleum feedstock through energy intensive processes involving toxic solvents and byproducts. The muscle protein titin, for example, endows muscle tissue with a combination of passive strength, damping capacity, and rapid mechanical recovery derived from titin’s UHMW ( >3 MDa) and highly repetitive sequence comprising hundreds of folded immunoglobulin (Ig) domains (Fig. 1a)[16,17,18] While these appealing mechanical properties have inspired many efforts to engineer titin-like materials[19,20,21,22,23], titin’s massive size and repetitive sequence have largely restricted these efforts to the production of titin-mimetic organic polymers rather than more environmentally-friendly protein-based materials (PBMs)[20]. We employ a synthetic biology approach to mitigate the challenges of genetic instability and low translational efficiency through in vivo protein polymerization catalyzed by split-inteins (SI) in Escherichia coli (Fig. 1b) In this manner, we microbially produce titin polymers with megadalton MW and subsequently develop an aqueous process to spin the resulting polymers into high-performance titin-based fibers that exhibit an intriguing combination of desirable mechanical properties
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