The ESTHER database, dedicated to ESTerases and alpha/beta-Hydrolase Enzymes and Relatives (https://bioweb.supagro.inra.fr/ESTHER/general?what=index), offers online access to a continuously updated, sequence-based classification of proteins harboring the alpha/beta hydrolase fold into families and subfamilies. In particular, the database proposes links to the sequences, structures, ligands and huge diversity of functions of these proteins, and to the related literature and other databases. Taking advantage of the promiscuity of enzymatic function, many engineered esterases, lipases, epoxide-hydrolases, haloalkane dehalogenases are used for biotechnological applications. Finding means for detoxifying those protein members that are targeted by insecticides, herbicides, antibiotics, or for reactivating human cholinesterases when inhibited by nerve gas, are still active areas of research. Using or improving the capacity of some enzymes to breakdown plastics with the aim to recycle valuable material and reduce waste is an emerging challenge. Most hydrolases in the superfamily are water-soluble and act on or are inhibited by small organic compounds, yet in a few subfamilies some members interact with other, unrelated proteins to modulate activity or trigger functional partnerships. Recent development in 3D structure prediction brought by AI-based programs now permits analysis of enzymatic mechanisms for a variety of hydrolases with no experimental 3D structure available. Finally, mutations in as many as 34 of the 120 human genes compiled in the database are now linked to genetic diseases, a feature fueling research on early detection, metabolic pathways, pharmacological treatment or enzyme replacement therapy. Here we review those developments in the database that took place over the latest decade and discuss potential new applications and recent and future expected research in the field.