Background: Nurses’ job satisfaction is a very important issue that determines the stability of the workforce, quality of treatment provided, and performance of a given organization. The level of demand in hospitals, staffing issues, and dependence on expatriate nurses in Saudi Arabia, especially in the Eastern Region, are growing, which is why the level of job satisfaction is crucial to assess. Aim: This study aimed to evaluate the extent of job satisfaction among nurses at major hospitals in the Eastern Region of Saudi Arabia and further determine institutional, job, and family related factors that affect job satisfaction based on Herzberg’s Two-Factor Theory. Methods: A cross-sectional quantitative design was used. A total of 390 registered nurses from four large hospitals were used as the data source using a structured self-administered questionnaire. The measures were institutional characteristics, job characteristics, family support, job satisfaction, absenteeism, and intention to leave. They were done using Descriptive statistics, independent t-tests, one-way ANOVA, Pearson correlation, and multiple regression were used. Results: The results showed moderate to high levels of overall job satisfaction among nurses. Job characteristics were found to be the best predictor of job satisfaction (b = 0.48, p < 0.001), followed by institutional characteristics (b = 0.34, p < 0.001) and family support (b = 0.16, p = 0.001). Major differences in job satisfaction were found in terms of age, experience, and clinical department. Conclusion: The Two-Factor Theory of Herzberg is in line with the findings that indicate that intrinsic job-related factors are the most determining factors in the job satisfaction of nurses in major hospitals in Eastern Saudi Arabia. Nurse satisfaction, retention, and quality of healthcare services can be enhanced through improvements in recognition, professional growth, and working environments.