Abstract
Tandem-repeat proteins comprise small secondary structure motifs that stack to form one-dimensional arrays with distinctive dynamic properties that are proposed to direct their cellular functions. Here, we report the force response of consensus-designed tetratricopeptide repeats (CTPRs) – superhelical arrays of short helix-turn-helix motifs. Not only are we able to directly observe the repeat-protein superhelix in the force data, but we also find that individual repeats undergo rapid fluctuations between folded and unfolded conformations resulting in a spring-like mechanical response. Using protein engineering, we show how the superhelical geometry can be altered by carefully placed amino-acid substitution and subsequently employ Ising model analysis to examine how these changes affect repeat stability and inter-repeat coupling in ensemble and single-molecule experiments. The Ising models furthermore allow us to map the unfolding pathway of CTPRs under mechanical load revealing how the repeat arrays are unzipped from both ends at the same time. Our findings provide the means to dissect and modulate repeat-protein dynamics and stability, thereby supporting ongoing research efforts into their functional relevance and exploiting them for nanotechnology and biomedical applications.
Competing Interest Statement
The authors have declared no competing interest.
