Modulating mechanical stability of heterodimerization between engineered orthogonal helical domains

Miao Yu,Zhihai Zhao,Shimin Le,Zibo Chen,Jie Yan

doi:10.1038/s41467-020-18323-w

Abstract

Mechanically stable specific heterodimerization between small protein domains have a wide scope of applications, from using as a molecular anchorage in single-molecule force spectroscopy studies of protein mechanics, to serving as force-bearing protein linker for modulation of mechanotransduction of cells, and potentially acting as a molecular crosslinker for functional materials. Here, we explore the possibility to develop heterodimerization system with a range of mechanical stability from a set of recently engineered helix-heterotetramers whose mechanical properties have yet to be characterized. We demonstrate this possibility using two randomly chosen helix-heterotetramers, showing that their mechanical properties can be modulated by changing the stretching geometry and the number of interacting helices. These helix-heterotetramers and their derivatives are sufficiently stable over physiological temperature range. Using it as mechanically stable anchorage, we demonstrate the applications in single-molecule manipulation studies of the temperature dependent unfolding and refolding of a titin immunoglobulin domain and α-actinin spectrin repeats.

Highlights

Stable specific heterodimerization between small protein domains have a wide scope of applications, from using as a molecular anchorage in single-molecule force spectroscopy studies of protein mechanics, to serving as force-bearing protein linker for modulation of mechanotransduction of cells, and potentially acting as a molecular crosslinker for functional materials
Their mechanical stability can be modulated by changing the force geometry and by changing the number of interacting helices
The mechanical stability of the helix-heterotetramers is greater under shearing force geometry than under unzipping force geometry, consistent with recent theoretical studies[40,54]

Summary

Introduction

Stable specific heterodimerization between small protein domains have a wide scope of applications, from using as a molecular anchorage in single-molecule force spectroscopy studies of protein mechanics, to serving as force-bearing protein linker for modulation of mechanotransduction of cells, and potentially acting as a molecular crosslinker for functional materials. We demonstrate this possibility using two randomly chosen helix-heterotetramers, showing that their mechanical properties can be modulated by changing the stretching geometry and the number of interacting helices These helix-heterotetramers and their derivatives are sufficiently stable over physiological temperature range. Using it as mechanically stable anchorage, we demonstrate the applications in single-molecule manipulation studies of the temperature dependent unfolding and refolding of a titin immunoglobulin domain and α-actinin spectrin repeats. The interactions between these helix-forming motifs, which result in formation of four-helix bundles (hereafter referred to as helixheterotetramers) with high thermal stability, are optimized for high-affinity orthogonal inter-helix interactions by computational models[43,48,49] (Fig. 1a) The orthogonality of these heterodimerization systems makes it appealing to develop a variety of mechanically stable heterodimerization systems from them. Whether these helix-heterotetramers have enough high mechanical stability over physiological temperature range is unclear

Methods

Results

Conclusion