AbstractPerformance fatigability is typically experienced as insufficient energy to complete daily physical tasks, particularly with advancing age, often progressing toward dependency. Thus, understanding the etiology of performance fatigability, especially cellular‐level biological mechanisms, may help to delay the onset of mobility disability. We hypothesized that skeletal muscle energetics may be important contributors to performance fatigability. Participants in the Study of Muscle, Mobility and Aging completed a usual‐paced 400‐m walk wearing a wrist‐worn ActiGraph GT9X to derive the Pittsburgh Performance Fatigability Index (PPFI, higher scores = more severe fatigability) that quantifies percent decline in individual cadence‐versus‐time trajectory from their maximal cadence. Complex I&II‐supported maximal oxidative phosphorylation (max OXPHOS) and complex I&II‐supported electron transfer system (max ETS) were quantified ex vivo using high‐resolution respirometry in permeabilized fiber bundles from vastus lateralis muscle biopsies. Maximal adenosine triphosphate production (ATP_max) was assessed in vivo by ³¹P magnetic resonance spectroscopy. We conducted tobit regressions to examine associations of max OXPHOS, max ETS, and ATP_max with PPFI, adjusting for technician/site, demographic characteristics, and total activity count over 7‐day free‐living among older adults (N = 795, 70–94 years, 58% women) with complete PPFI scores and ≥1 energetics measure. Median PPFI score was 1.4% [25th–75th percentile: 0%–2.9%]. After full adjustment, each 1 standard deviation lower max OXPHOS, max ETS, and ATP_max were associated with 0.55 (95% CI: 0.26–0.84), 0.39 (95% CI: 0.09–0.70), and 0.54 (95% CI: 0.27–0.81) higher PPFI score, respectively. Our findings suggested that therapeutics targeting muscle energetics may potentially mitigate fatigability and lessen susceptibility to disability among older adults.