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Mechanical stability of bonds and protein interactions has recently become accessible through single molecule mechanical experiments. So far, mechanical information about molecular bond mechanics has been largely limited to a single direction of force application. However, mechanical force acts as a vector in space and hence mechanical stability should depend on the direction of force application. In skeletal muscle, the giant protein titin is anchored in the Z-disk by telethonin. Much of the structural integrity of the Z-disk hinges upon the titin-telethonin bond. In this paper we show that the complex between the muscle proteins titin and telethonin forms a highly directed molecular bond. It is designed to resist ultra-high forces if they are applied in the direction along which it is loaded under physiological conditions, while it breaks easily along other directions. Highly directed molecular bonds match in an ideal way the requirements of tissues subject to mechanical stress.

Single-molecule methods have given experimental access to the mechanical properties of single protein molecules. So far, access has been limited to mostly one spatial direction of force application. Here, we report single-molecule experiments that explore the mechanical properties of a folded protein structure in precisely controlled directions by applying force to selected amino acid pairs. We investigated the deformation response of GFP in five selected directions. We found fracture forces widely varying from 100 pN up to 600 pN. We show that straining the GFP structure in one of the five directions induces partial fracture of the protein into a half-folded intermediate structure. From potential widths we estimated directional spring constants of the GFP structure and found values ranging from 1 N/m up to 17 N/m. Our results show that classical continuum mechanics and simple mechanistic models fail to describe the complex mechanics of the GFP protein structure and offer insights into the mechanical design of protein materials.