Misalignment is one of the key reasons for vibrations in most of the rotating system. The present study focuses on interactions among speed, load, and defect severity by investigating their effect on the system vibration. Response surface methodology (RSM) with root-mean-square (RMS) as a response factor is used to understand the influence of such interactions on the system performance. Experiments are planned using design of experiments, and analysis is carried out using analysis of variance (ANOVA). It is observed that speed has a remarkable effect on RMS value in both parallel and angular types of misalignment and affects the system performance. RSM results revealed that a change in load has less impact on vibration amplitude in case of horizontal and vertical directions, but there is a significant variation in RMS value in axial direction for both types of misalignment. A slight increase in the RMS value with an increase in defect severity is observed in the axial direction. These observations will help to understand the misalignment defect and its effect in a better way.