In response to physical exercise and diet, skeletal muscle adapts to energetic demands through large transcriptional changes. This remodelling is associated with changes in skeletal muscle DNA methylation which may participate in the metabolic adaptation to extracellular stimuli. Yet, the mechanisms by which muscle-borne DNA methylation machinery responds to diet and exercise and impacts muscle function are unknown. Here, we investigated the function of de novo DNA methylation in fully differentiated skeletal muscle. We generated muscle-specific DNA methyltransferase 3A (DNMT3A) knockout mice (mD3AKO) and investigated the impact of DNMT3A ablation on skeletal muscle DNA methylation, exercise capacity and energy metabolism. Loss of DNMT3A reduced DNA methylation in skeletal muscle over multiple genomic contexts and altered the transcription of genes known to be influenced by DNA methylation, but did not affect exercise capacity and whole-body energy metabolism compared to wild type mice. Loss of DNMT3A did not alter skeletal muscle mitochondrial function or the transcriptional response to exercise however did influence the expression of genes involved in muscle development. These data suggest that DNMT3A does not have a large role in the function of mature skeletal muscle although a role in muscle development and differentiation is likely.