The aim of this study was to assess the relationships between physical activity level and anxiety, depression, and functional ability in children and adolescents with juvenile idiopathic arthritis (JIA). Cross-sectional study design including patients with JIA aged between 8 and 17 years and healthy controls was used. Sociodemographic data and clinical features were assessed. Physical activity level and energy expenditure were assessed with a 1-day activity diary. Anxiety was screened by The Screen for Child Anxiety Related Emotional Disorders (SCARED) questionnaire. Depressive symptoms were assessed by the Children's Depression Inventory (CDI). Functional ability was assessed with the Childhood Health Assessment Questionnaire (CHAQ). Pain and overall well-being were measured using a visual analog scale (VAS). Fifty-two patients and 48 controls were included with a mean age of 12.13 ± 2.92 and 11.27 ± 1.59 years, respectively. The mean disease duration was 64 months. The JIA group had significantly less time in physical activity (p = 0.000), decrease in energy expenditure (p = 0.04), and higher CHAQ scores (p = 0.000) compared with the control group. In the JIA group, significant relationships were found between the number of active joint and disease duration (r = 0.44, p = 0.000) and VAS pain (r = 0.30, p = 0.02), between SCARED and CDI (r = 0.54, p = 0.000). Significant relationships were found between VAS overall well-being and CDI (r = 0.29, p = 0.03), CHAQ (r = 0.37, p = 0.000), and VAS pain (r = 0.41, p = 0.000). Correlation between CHAQ and CDI (r = 0.34, p = 0.01) was significant. The result of our study suggested that only depression was related to anxiety, functional ability, and well-being in children and adolescents with JIA.