Integrins mediate cell adhesion to extracellular matrix and transduce signals bidirectionally across the membrane. Integrin α_Vβ₃ has been shown to play an essential role in tumor metastasis, angiogenesis, hemostasis and phagocytosis. Integrins can take several conformations, including the bent and extended conformations of the ectodomain, which regulate integrin functions. Using a biomembrane force probe, we characterized the bending and unbending conformational changes of single α_Vβ₃ integrins on living cell surfaces in real-time. We measured the probabilities of conformational changes, rates and speeds of conformational transitions, and the dynamic equilibrium between the two conformations, which were regulated by tensile force, dependent on the ligand, and altered by point mutations. These findings provide insights into how α_Vβ₃ acts as a molecular machine and how its physiological function and molecular structure are coupled at the single‐molecule level.