The aim of this study was to investigate the effect of fatigue on electromyographic (EMG) parameters of healthy young adults during obstacle crossing of two different heights. Twelve untrained male adults (23 ± 5 years of age) were fatigued running on a treadmill with increasing speed and inclination and walked over an obstacle with a height set at 10% and 20% of each individual’s lower limb length. Maximal plantar flexor torque and EMG of the medial gastrocnemius, soleus, and tibialis anterior muscles of the trailing limb were assessed during obstacle crossing. Data were captured before, immediately after and 5 minutes after a fatigue session. Fatigue induced significant reduction on the plantar flexor torque output immediately after and 5 minutes after exhaustion. After fatigue gait speed was not affected, the minimum distance between the obstacle and the trailing or leading foot remained unchanged, and the trailing foot contacted the ground closer to the obstacle immediately after fatigue. Regarding the EMG, medial gastrocnemius became after fatigue more active during swing phase when increasing the obstacle height, whereas this was not the case before or 5 minutes after fatigue. No other significant difference was observed for any of the examined muscles. It is concluded that the assessed fatigue protocol induced only minimal changes in the EMG activity of the examined muscles during obstacle crossing. Therefore, it is suggested that the neuromuscular system of healthy young individuals is able to respond to the decreased force capacity after fatigue during obstacle crossing of heights up to the 20% of the limb length.