Introduction: Proton-induced nucleon knockout reactions have been widely used in recent years to explore the single-particle properties of various nuclei. In this study, we examine the effects of the single-particle overlap function on the cross sections and spectroscopic factors of the 16 O(p,2p) 15 N reactions. Methods: These effects are extensively studied through an analysis of the cross sections of the 16 O(p,2p) 15 N reaction at incident energies of 392 and 505 MeV. The evaluation is carried out within the framework of the distorted-wave impulse approximation (DWIA), utilizing overlap function inputs from different structure calculations and potential prescriptions. Results: The analysis reveals that for the 16 O(p,2p) 15 N reaction and the chosen incident energies, the ambiguities arising from different selections of the overlap function are small but nonnegligible. Conclusion: Given the existence of these theoretical uncertainties, it is important that careful and consistent choices of single-particle overlap functions are made in systematic knockout reaction analyses to ensure the extraction of reliable structure information.