Motivational factors play an important role in increasing employee job satisfaction. Satisfied employees in return can help in improving organizational performance. The objective of this study is to analyze the effects of motivational factors on employee job satisfaction. This is an exploratory study based on primary data. The primary data has been collected from non-academic staff of University of the Punjab, which is one of the biggest universities of Pakistan. The study has tested Herzberg et al.’s (1959) well-known, duality theory of motivators and hygiene factors and the impact of personal characteristics and job characteristics on perceptions of the work environment and job satisfaction. Structural equation modeling technique has been applied to test hypothesis, SPSS 16.0 has also been adopted for basic analysis purposes. The results offer inconclusive support of Herzberg’s theory. The study concludes that intrinsic motivational factors are having significant relationship with employee job satisfaction, whereas hygiene (extrinsic) factors are not having any significant relationship with employee job satisfaction. Moreover, significant difference was observed between gender, qualification, experience, job characteristics and job satisfaction. This study offers useful information by discussing both practical implications for those in leadership positions in Public Sector University set up in Pakistan, and theoretical implications for researchers interested in exploring job satisfaction in a higher education context.