Multidomain proteins under force

Abstract

Advancements in single molecule force spectroscopy techniques such as atomic force microscopy and magnetic tweezers allow investigating how domain folding under force can have physiological roles. Combining these techniques with protein engineering and HaloTag covalent attachment, we investigate similarities and differences between four model proteins: I10 and I91 – two immunoglobulin-like domains from the muscle protein titin, and two α+β fold proteins – ubiquitin and protein L. These proteins show a different mechanical response and have unique extensions under force. Remarkably, when normalized to their contour length, the size of the unfolding and refolding steps as a function of force reduces to a single master curve. This curve can be described using standard polymer elasticity models, explaining the entropic nature of the measured steps. We further validate our measurements with a simple energy landscape model, which combines protein folding with polymer physics and accounts for the complex nature of tandem domains under force. This model can become a useful tool toward deciphering the complexity of multidomain proteins operating under force.

Chemical unfolding of protein domains induces shape change in programmed protein hydrogels

“…Using this approach, we determine the Young's modulus from the initial slope of each stress-strain curve, which reports on the gel stiffness. Furthermore, as proteins unfold and refold at vastly different forces 23 , the stress-strain curves show important hysteresis, which reports on the energy being dissipated due to these phase transitions 19,24 .…”

Proteins Breaking Bad: A Free Energy Perspective

“…From a more global perspective, the observations described here provide significant evidence for an element often dismissed when discussing protein folding under force: the polymer properties of the polypeptide chain. As we have shown, the end-to-end distance of the unfolded protein scales with force following polymer models such as the freely-jointed chain, which is inherently nonlinear, and acquires over half of its maximum extension below 10 pN, just in the range where proteins fold 33 . This fact is surprisingly dodged in most descriptions on protein folding/unfolding under force, starting with the fact that force is often modeled as an effective tilt in the landscape with a - F·x term 36–39 .…”

“…Additionally, the polymer character of the polypeptide chain together with protein hydrophobic collapse modulate the folding properties of polyproteins under force, since the landscape is tilted with non-linear dependence ( U FJC –F·x ). Interestingly, most proteins fail to fold above 10 pN, and exhibit and abrupt decrease in the folding probability in a range of few picoNewton, in general from 6 – 10 pN 2, 5, 33, 43–44 . Our model is able to capture this behavior, via the force dependency of the global minimum, whose force sensitivity is well understood considering the steepness induced by the Morse potential and the non-linearity of the freely-jointed chain model.…”

Mechanical Deformation Accelerates Protein Ageing

“…The advent of ultra-stable magnetic tweezers instrumentation now permits studies of protein dynamics under force over extended time periods. [9] Here, we use magnetic tweezers to monitor the folding dynamics of single proteins placed under force, over time scales of hours to days, and study how they age. We expose single proteins to the cumulative oxidative modifications of cryptic side chains, and study the effect on elasticity.…”

“…Am ajor difficulty in the study of oxidative protein damage is the extremely long time scales involved and the heterogeneity of this chemistry.Given that most proteins are only transiently unfolded in vivo,the probability of oxidative modification of cryptic side chains is low.N evertheless, damage accumulates over time,p articularly in low-turnover proteins.T he advent of ultra-stable magnetic tweezers now permits studies of protein dynamics under force over extended time periods. [9] Here,w eu se magnetic tweezers to monitor the folding dynamics of single proteins placed under force,o ver time scales of hours to days,a nd study how they age.W ee xpose single proteins to the cumulative oxidative modifications of cryptic side chains,a nd study the effect on elasticity.I nd oing so,w es how that keeping ap rotein in the unfolded state by applying force is af orm of accelerated ageing, where decades worth of oxidative damage are compressed into hours.This ageing causes aloss of elasticity,with aged proteins providing 50 %l ess contractility than younger proteins.…”