Cardiorespiratory fitness is linked to various health outcomes. This study aimed to establish cut-off values for estimated cardiorespiratory fitness associated with poor physical functioning in middle-aged to older adults. A total of 2638 participants (1083 males), aged 40-69 years, performed a 6-min walking test (6MWT) to estimate cardiorespiratory fitness (VO<inf>2max)</inf> and completed a health-related questionnaire. Physical functioning was assessed using four outcomes: perceived health, perceived fitness, difficulties in 2-km walking, and difficulties in climbing several stairs. Receiver operator characteristics (ROC) curves were used to determine cut-off values for estimated VO<inf>2max</inf> and 6MWT distance linked to poor physical functioning based on the four outcomes. Analyses were done separately for males and females, and for 40- to 49-, 50- to 59-, and 60- to 69-year-old groups. Multinomial regression was used to analyze differences between fitness thirds in outcome variables. ROC analyses were significant for all outcomes (AUC 0.686-0.834, P<0.001). In the total group, cut-off values for estimated VO<inf>2max</inf> were 29.9-30.2 mL/kg/min, and for walking distance, 588.0-599.5 m. Cut-off values decreased with age, and mean differences between males and females were 3.3 mL/kg/min and 33 m. All cut-off values were within ±1.3 mL/kg/min of the lower fitness tertile. The odds ratios of having poor physical functioning in the middle and high fitness thirds were less than 0.50 compared to low fitness third. VO<inf>2max</inf> of 30 mL/kg/min and 600-m walking distance seemed reasonable cut-off values for poor physical functioning. Exceeding the currently proposed thresholds is a desirable goal to avoid consequences of insufficient fitness.