Titin as a mechanical damper: Balancing Stability and longevity through inter-domain linker design

Abstract

Titin, a giant protein ([~]3-4 MDa), functions as a molecular-spring to regulate muscle elasticity. More than 90% of Titin is composed of domains that absorb mechanical energy and undergo stochastic unfolding-refolding under tension ([~]tens of pN). These domains are connected in tandem by interdomain linkers (IDLs), which constitute less than 10% of the total mass. Despite their small genomic footprint, bioinformatics mapping suggests that IDLs have an outsized impact on protein mechanics, potentially contributing to disease pathology. Using magnetic tweezers, here we examine how linkers influence mechano-response of domains to constant and oscillatory forces. We found that short linkers limit interdomain movement and promote first-order cooperative folding transitions of domains. In contrast, long flexible linkers induce creep-like deformations interspersed with sharp, stepwise transitions. Surprisingly, linkers that improve domain-stability resist unfolding under constant pulling forces, but lose power retention faster under oscillatory forces. Our findings reveal a trade-off between mechanical stability and energy retention in titin, a key muscle protein. These insights offer new design principles for mechano-responsive protein engineering. TeaserTiny linkers fine-tune how bulky domains in titin respond to force.