Haloacid dehalogenases (HAD) are members of a large superfamily that includes many Structural Genomics proteins with poorly characterized functionality. This superfamily consists of multiple types of enzymes that can act as sugar phosphatases, haloacid dehalogenases, phosphonoacetaldehyde hydrolases, ATPases, or phosphate monoesterases. Here, we report on predicted functional annotations and experimental testing by direct biochemical assay for Structural Genomics proteins from the HAD superfamily. To characterize the functions of HAD superfamily members, nine representative HAD proteins and 21 structural genomics proteins are analyzed. Using techniques based on computed chemical and electrostatic properties of individual amino acids, the functions of five structural genomics proteins from the HAD superfamily are predicted and validated by biochemical assays. A dehalogenase-like hydrolase, RSc1362 (Uniprot Q8XZN3, PDB 3UMB) is predicted to be a dehalogenase and dehalogenase activity is confirmed experimentally. Four proteins predicted to be sugar phosphatases are characterized as follows: a sugar phosphatase from Thermophilus volcanium (Uniprot Q978Y6) with trehalose-6-phosphate phosphatase and fructose-6-phosphate phosphatase activity; haloacid dehalogenase-like hydrolase from Bacteroides thetaiotaomicron (Uniprot Q8A2F3; PDB 3NIW) with fructose-6-phosphate phosphatase and sucrose-6-phosphate phosphatase activity; putative phosphatase from Eubacterium rectale (Uniprot D0VWU2; PDB 3DAO) as a sucrose-6-phosphate phosphatase; and hypothetical protein from Geobacillus kaustophilus (Uniprot Q5L139; PDB 2PQ0) as a fructose-6-phosphate phosphatase. Most of these sugar phosphatases showed some substrate promiscuity.
Functional annotation of haloacid dehalogenase superfamily structural genomics proteins
Lakindu S. Pathira Kankanamge, Lydia A. Ruffner, Mong Mary Touch, Manuel Pina, Penny J. Beuning, Mary Jo Ondrechen; Functional annotation of haloacid dehalogenase superfamily structural genomics proteins. Biochem J 11 October 2023; 480 (19): 1553–1569. doi: https://doi.org/10.1042/BCJ20230057
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