The haloacid dehydrogenase (HAD) superfamily is an evolutionarily diverse, widely distributed, and abundant class of enzymes (Koonin and Tatusov, 1994). Within this superfamily, phosphatases, which cleave CO–P bonds, are the most abundant, accounting for approximately 79 % of the enzymes (Kuznetsova et al., 2015). These are followed by ATPases (cleave PO–P bonds), phosphoesterases (cleave CP bonds), dehalogenases (cleave CCl bonds), and phosphoglucose mutases (cleave/transfer CO–P bonds) (Koonin and Tatusov, 1994; Meng et al., 2019). To date, the InterPro database lists over one million protein sequences of HAD enzymes (IPR023214); however, in only approximately 20 % of these sequences have catalytic function been annotated. Numerous types of HAD enzymes are found in both prokaryotes and eukaryotes, including at least 23 types identified in Escherichia coli and at least 19 types found in yeast (Kuznetsova et al., 2015; Kuznetsova et al., 2006). In summary, the diverse HAD enzymes provide a valuable reservoir of enzymes for biocatalytic processes involving phosphatases.

Phosphatases, which remove phosphate groups from biomolecules, can be classified into several types according to their substrates, including protein phosphatases (Brautigan, 2013; Tonks, 2013), sugar phosphatases (Lu et al., 2005), lipid phosphatases (McDermott et al., 2006), and nucleotide phosphatases (Kuipers et al., 2016). Phosphatases are essential for various cellular functions, including the maintenance of metabolic pools, regulation of primary and secondary metabolism, cellular housekeeping, and nutrient absorption (Kuznetsova et al., 2006; Meng et al., 2019; Rangarajan et al., 2006). Phosphatases are known for their substrate promiscuity, meaning that they interact with a broad range of substrates. Huang et al. constructed a substrate library containing 167 phosphorylated compounds and analyzed the substrate profiles of over 217 phosphatases. The results showed that more than 75 % of these phosphatases exhibited dephosphorization activity against at least five substrates (Huang et al., 2015). Phosphatases with a broad scope of substrates show promise as tools to enhance cellular metabolism by allowing the evolution of new enzyme activity in response to environmental challenges, reducing the number of phosphatase genes required for dephosphorylation reactions across various substrates and increasing evolutionary flexibility (Huang et al., 2015; Meng et al., 2019).

The abundance of phosphatases provides a wide range of multifunctional candidate enzymes for the dephosphorylation of metabolites containing phosphate groups, which makes them suitable for various applications, such as biomanufacture of high-value compounds (You and Percival Zhang, 2017), drug discovery (Köhn, 2020; Mullard, 2018), biomarker and biosensor development (Han et al., 2020a), and environmental remediation (Wang et al., 2021). In particular, the irreversible and thermodynamically favorable dephosphorization reaction catalyzed by sugar phosphatase enables in vitro synthetic enzymatic biosystems (ivSEBS) to achieve high conversion rates from substrate to product, making it feasible for the production of various high-value functional sugars (Meng et al., 2019; Zhang et al., 2023). However, in the industrial application of enzymatic catalysis, the substrate promiscuity of phosphatases can lead to the undesired dephosphorization of intermediates within the ivSEBS. This results in the formation of by-products and reduced substrate conversion rates, which significantly impede the industrialization of ivSEBS (Meng et al., 2019). Furthermore, the substrate profile of a large number of phosphatases remains poorly studied, posing a significant challenge in identifying enzymes with strong specificity to a particular substrate. Additionally, the poor correlation between the sequence, structure, and function of phosphatases makes substrate specificity prediction difficult in many cases (Fahs et al., 2016).

Recent advancements in sugar phosphatase research have significantly enhanced our understanding and applications of these enzymes. In this review, we present the latest progress in this field and offer a thorough analysis of sugar phosphatase classification, structural characteristics, substrate recognition mechanisms, and molecular modifications. We also provide an in-depth examination of the enzymatic synthesis of functional sugars using sugar phosphatase-driven in vitro enzymatic biosystems. Lastly, guidelines for identify sugar phosphatase towards desired substrate are recommended. The overall objective is to provide a comprehensive understanding of sugar phosphatases and to inspire further developments in protein engineering and enzymatic synthesis of high-value compounds via these promising enzymes.