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. 2021 Aug 30;12:5182. doi: 10.1038/s41467-021-25360-6

Fig. 1. The multi-scale structure of muscle and schematic representation of SI-based polymerization of the titin protein in E. coli.

Fig. 1

a Muscle tissue (1) is composed of specialized, elongated (> 1 cm) cells called muscle fibers (2). Muscle fibers are packed with proteinaceous myofibrils (3) that span the entire length of the cell. Myofibrils are composed of repeating stacks of chemically controllable, contractile elements called sarcomeres. (4) Sarcomeres are composed primarily of three proteins: actin, myosin, and titin. Titin spans half the length of the sarcomere, anchoring the opposing Z- and M-lines, and consists of hundreds of repeating immunoglobulin (Ig) domains that are integral to the passive strength (i.e., resistance to deformation without energy input), damping capacity, and mechanical recovery of the macroscopic muscle fiber. The images of muscle fiber and myofibril are modified from the OpenStax Anatomy and Physiology Textbook Version 8.25, Published May 18, 2016 (OpenStax, CC BY 4.0 https://creativecommons.org/licenses/by/4.0) and the image of the sarcomere is modified from Giganti, D., Yan, K., Badilla, C.L. et al. Disulfide isomerization reactions in titin immunoglobulin domains enable a mode of protein elasticity. Nat Commun 9, 185 (2018). 10.1038/s41467-017-02528-7, (David Giganti, Kevin Yan, Carmen L. Badilla, Julio M. Fernandez & Jorge Alegre-Cebollada, CC BY 4.0 https://creativecommons.org/licenses/by/4.0). b To facilitate the production of UHMW titin polymer in vivo, a relatively small, genetically stable, 41.3 kDa rabbit soleus titin protein-coding sequence (4 Ig; purple) was flanked by complimentary SIs, gp41-1C (IntC; green) and gp41-1N (IntN; blue) (i). DNA sequence-recoded IntC-4Ig-IntN was produced in an engineered E. coli host under the control of inducible promoter PLacO-1 (ii). The SI-flanked monomer protein was overexpressed in bioreactor cultures (iii) and polymerized intracellularly through successive rounds of SI-catalyzed intermolecular ligation to produce UHMW titin (iv). Purification and processing yielded microbially produced titin fibers that recapture the damping capacity and mechanical recovery of muscle along with high strength and toughness (v). Titin 4 Ig and SI structures were acquired using PBD accession numbers 3B43 and 6QAZ, respectively.