Soluble sugar and organic acids are key determinants of fruit organoleptic quality and directly affect the commodity value and economic returns of fruit crops. We performed whole-genome sequencing of the apple varieties Gala and Xiahongrou, along with their F1 hybrids, to construct a high-density bin map. Our quantitative genetic analysis pinpointed 53 quantitative trait loci (QTLs) related to 11 sugar and acid traits. We identified a candidate gene, MdNADP-ME, responsible for malate degradation, in a stable QTL on linkage group 15. Sequence analysis revealed an A/C SNP in the promoter region (MEp-799) that influences binding of the MdMYB2 transcription factor, thereby affecting MdNADP-ME expression. In our study of various apple genotypes, this SNP has been demonstrated to be linked to malate and fructose levels. We also developed a dCAPS marker associated with fruit fructose content. These results substantiate the role of MdNADP-ME in maintaining the equilibrium between sugar and acid contents in apple fruits.