Anisotropy in mechanical unfolding of protein upon partner-assisted pulling and handle-assisted pulling

Abstract

Proteins as force-sensors respond to mechanical cues and regulate signaling in physiology. Proteins commonly connect the source and response points of mechanical cues in two conformations, independent proteins in end-to-end geometry and protein complexes in handshake geometry. The force-responsive property of independent proteins in end-to-end geometry is studied extensively using single-molecule force spectroscopy (SMFS). The physiological significance of the complex conformations in force-sensing is often disregarded as mere surge protectors. However, with the potential of force-steering, protein complexes possess a distinct mechano-responsive property over individual force-sensors. To decipher, we choose a force-sensing protein, cadherin-23, from tip-link complex and perform SMFS using end-to-end geometry and handshake complex geometry. We measure higher force-resilience of cadherin-23 with preferential shorter extensions in handshake mode of pulling over the direct mode. The handshake geometry drives the force-response of cadherin-23 through different potential-energy landscapes than direct pulling. Analysis of the dynamic network structure of cadherin-23 under tension indicates narrow force-distributions among residues in cadherin-23 in direct pulling, resulting in low force-dissipation paths and low resilience to force. Overall, the distinct and superior mechanical responses of cadherin-23 in handshake geometry than single protein geometry highlight a probable evolutionary drive of protein-protein complexes as force-conveyors over independent ones.

Tip-links serve as force-pass filter to fulfil the role of gating-springs

“…linker between EC2-3 domains, there also we observed that the linker is pre-compressed (Figure S9a) . Further, the ∼12 nm extension, referring as partial domain unfolding(Arora et al, 2021), may diffuse force non-elastically and could be the dashpots in the spring. Notably, a gating-spring by definition, must possess elastic and non-elastic (dashpot) compliances.…”

“…Dynamic alterations in the entropic conformations of the extraordinarily long tip-links cadherins under tension are already reported (Arora et al, 2021;Bartsch et al, 2018;Choudhary et al, 2020;Oroz et al, 2019). How such alterations contribute to the bond-lifetime dynamics of tip-links is not yet clear.…”

“…In addition, we verified the effect of mutation associated with congenital hearing loss on the bond-lifetime dynamics. Moreover, the existing biophysical, genetic screening in model systems and other physiological studies verify that Cdh23 may be the dominant elastic component of tiplinks (Arora et al, 2021;Astuto et al, 2002;Miyagawa et al, 2012). Here, we used different variants of Cdh23 to understand how alterations in the protein elasticity affect the bond-lifetime dynamics of tip-links.…”

“…Kinks and bents in proteins are mechanical linkages that contribute to the elasticity of the tertiary structure(Granzier et al, 1997; Lee et al, 2006). Accordingly, individual tip-links proteins and the tip-links complex too can exploit their tertiary structure elasticity in response to mechanical stimuli and contribute to force-dissipation(Arora et al, 2021; Bartsch et al, 2018; Choudhary et al, 2020; Oroz et al, 2019). In fact, the mechanical origin of force-dissipation for individual tip-links is previously identified from non-equilibrium simulations and experiments (Arora et al, Choudhary et al, 2020), however, their impact on the tip-links complex interface is still elusive.…”

“…However, it demands further clarity in the functional model to establish tip-links as gating-springs.Traditionally, temporal response to constant force or stress has been the classical physics experiment to study the viscoelasticity of soft matters, extrapolation to which provided the spring models. Notably, several pulling experiments have already been done with individual tip-links proteins which deciphered the axial elasticity of the proteins (Arora et al, 2021;Bartsch et al, 2018;Oroz et al, 2019). While these studies quantitatively identify and characterize the elastic and non-elastic components of tip-links proteins, they failed to identify the characteristic features of a gating-spring.…”