Understanding the three-dimensional structure and mechanical response of biomolecules is key to uncovering their molecular mechanisms, particularly in contexts where force plays a regulatory role. Structural methods such as X-ray crystallography, cryo-electron microscopy, and NMR spectroscopy provide high-resolution conformational data, while single-molecule force spectroscopy reveals mechanical properties, but these approaches are rarely integrated. A more comprehensive understanding of structure-function relationships, including non-equilibrium conformations and transitions under force, calls for methods capable of simultaneously resolving structural and mechanical properties at the single-molecule level. To meet this need, we present a DNA nanoswitch calipers platform capable of both measuring multiple intramolecular distances and mechanically unfolding individual biomolecules along defined axes. Using human telomeric DNA G-quadruplexes as a model system, we mapped distances between labeled sites to distinguish conformational states and performed directional unfolding to characterize mechanical stability along defined axes. This integrative approach revealed subtle conformational and mechanical differences, showcasing DNA nanoswitch calipers as a modular, broadly applicable approach for mechanostructural analysis of complex biomolecular systems.