Protein dynamics has been recognized as a factor responsible for the powerful catalysis. Over the decades, many experimental and theoretical studies have provided the evidence that dynamic fluctuation positively contributed to their catalysis; however, how the fluctuation affect which elementary step of catalysis remains poorly understood. Recently, in order to elucidate at elementary-step resolution, various proteins are observed at the single-molecule level. Among them F1-ATPase (F1) is the excellent system in which we can not only measure almost all elementary-steps of the catalysis, but also apply the indicate amount of the viscous load on its conformational change. Accordingly, we tried to characterize the effect of the dynamic fluctuation on the catalysis of F1. F1 is the rotary molecular motor, which couples ATP hydrolysis to rotary motion. F1 consists of α3β3γ subunits, in which α3β3 and γ compose the stator ring and the rotary shaft. In order to visualize the rotary motion, we attached the rotary probe on γ. The size of the probe was much bigger than that of γ, and therefore, the actual viscous drag of γ was increased, in other word, thermal fluctuation became slower. Then, we observed the rotary motion by using the various sizes of probes (f=40∼500nm), and studied the effect of the dynamics of the attached probes on the rate of ATP hydrolysis at elementary-step resolution. The rates of ATP binding and cleavage slightly depended on the viscous drag; on the other hand, Pi releasing rate was drastically accelerated as the viscous drag was decreased. Furthermore, we build the simulation model based on the previous study, and well reproduced and confirmed the experimental result computationally. This study is the achievement in which the effect of the thermal fluctuation on the catalysis is quantitatively evaluated at elementary-step resolution.