Mental fatigue is one of the main reasons for the decline of response inhibition. This study aimed to explore the impairing influence of mental fatigue on a driver's response inhibition. The effects of mental fatigue on response inhibition were assessed by comparing brain activity and behavioral indices when performing a Go/NoGo task before and after a 90-min fatigue manipulation task. Participants in the driving group performed a simulated driving task, while individuals in the control group spent the same time watching movies. We found that participants in the driving group reported higher levels of mental fatigue and had a higher percentage of eye closure and larger lateral deviations from their lane positions, which indicated there was effective manipulation of mental fatigue through a prolonged simulated driving task. After manipulation of mental fatigue, we observed increased reaction time and miss rates, delayed NoGo-N2 latency and Go-P3 latency, and decreased NoGoP3 amplitude, which indicated that mental fatigue may slow down the speed of the inhibition process, delay the evaluation of visual stimuli and reduce the availability of attentional resources. These findings revealed the underlying neurological mechanisms of how mental fatigue impaired response inhibition.