Hexokinase (HXK) is the first irreversible catalytic enzyme in the glycolytic pathway, which not only provides energy for plant growth and development but also serves as a signaling molecule in response to environmental changes. However, the evolutionary pattern of the HXK gene family in apple remains unknown. In this study, a total of nine HXK genes were identified in the Malus×domestica genome GDDH13 v1.1. The physiological and biochemical properties, exonintron structures, conserved motifs, and cis-elements of the MdHXK genes were determined. Predicted subcellular localization indicated that the MdHXK genes were mainly distributed in the mitochondria, cytoplasm, and nucleus. Gene duplication revealed that whole-genome duplication (WGD) and segmental duplication played vital roles in MdHXK gene family expansion. The ω values of pairwise MdHXK genes indicated that this family was subjected to strong purifying selection during apple domestication. Additionally, five subfamilies were classified, and recent/old duplication events were identified based on phylogenetic tree analysis. Different evolutionary rates were estimated among the various HXK subfamilies. Moreover, divergent expression patterns of the MdHXK genes in four source-sink tissues and at five different apple fruit developmental stages indicated that they play vital roles in apple fruit development and sugar accumulation. Our study provides a theoretical basis for future elucidation of the biological functions of the MdHXK genes during apple fruit development.

己糖激酶（HXK）是糖酵解途径中第一个不可逆的催化酶，它不仅为植物生长发育提供能量，而且作为响应环境变化的信号分子。然而，苹果中HXK基因家族的进化模式仍然未知。在这项研究中，共有九个HXK基因进行鉴定平邑×家蝇基因组GDDH13 V1.1。测定了MdHXK基因的生理和生化特性、外显子结构、保守基序和顺式元件。预测的亚细胞定位表明MdHXK基因主要分布在线粒体、细胞质和细胞核中。基因重复显示全基因组重复（WGD）和节段重复在MdHXK基因家族扩展中起着至关重要的作用。成对MdHXK基因的 ω 值表明该家族在苹果驯化过程中受到强烈的纯化选择。此外，对五个亚科进行了分类，并根据系统发育树分析确定了最近/旧的重复事件。在不同的 HXK 亚科中估计了不同的进化率。此外，MdHXK 的不同表达模式四个源库组织和五个不同苹果果实发育阶段的基因表明它们在苹果果实发育和糖分积累中起着至关重要的作用。我们的研究为未来阐明MdHXK基因在苹果果实发育过程中的生物学功能提供了理论基础。