The emergency department (ED) is a stressful workplace for nurses, due to heavy workload and work shifts, which affect sleep quality, causing occupational fatigue. As a result, nurses burn out and turn over and quality of care is impaired. The aim of this study was to investigate sleep disturbance (SD) and occupational fatigue (OF) in emergency nurses of public hospitals in Greece. This descriptive cross-sectional study was conducted in the ED of tertiary public hospitals in Greece from September 2019 to October 2021, after the approval from the scientific and ethical committee of hospitals and the full informed consent of participants. The Standard Shift work Index (SSI) questionnaire was used, which includes 12 question groups about sleep habits and OF evaluation for shift workers, as nurses. Two hundred and ten ED nurses were recruited for the purpose of the study. In this research their quality of life, sleep completeness, and fatigue levels were studied. A moderate level of SD (total score = 70.54) of nurses was found that was related to marital status, presence of children, and underlying diseases (p = 0.012, p = 0.024, and p = 0.002, respectively). OF was assessed at low levels (score = 27.34) and was mainly related to age, with younger nurses reporting less fatigue compared with older nurses. The effect of work shifts in SD and OF of ED nurses was well documented. ED nurses' OF was correlated with increased SD, age, and marital status. Further research is needed for a better assessment and understanding of the factors that influence SD and OF of ED nurses to plan a strategy to optimize sleep patterns of work shift ED nurses.