Abstract
We characterized Teth514_1788 and Teth514_1789, belonging to glycoside hydrolase family 130, from Thermoanaerobacter sp. X-514. These two enzymes catalyzed the synthesis of 1,2-β-oligomannan using β-1,2-mannobiose and d-mannose as the optimal acceptors, respectively, in the presence of the donor α-d-mannose 1-phosphate. Kinetic analysis of the phosphorolytic reaction toward 1,2-β-oligomannan revealed that these enzymes followed a typical sequential Bi Bi mechanism. The kinetic parameters of the phosphorolysis of 1,2-β-oligomannan indicate that Teth514_1788 and Teth514_1789 prefer 1,2-β-oligomannans containing a DP ≥3 and β-1,2-Man2, respectively. These results indicate that the two enzymes are novel inverting phosphorylases that exhibit distinct chain-length specificities toward 1,2-β-oligomannan. Here, we propose 1,2-β-oligomannan:phosphate α-d-mannosyltransferase as the systematic name and 1,2-β-oligomannan phosphorylase as the short name for Teth514_1788 and β-1,2-mannobiose:phosphate α-d-mannosyltransferase as the systematic name and β-1,2-mannobiose phosphorylase as the short name for Teth514_1789.
Highlights
Glycoside phosphorylases catalyze the cleavage of glycosyl linkages via a substitution with inorganic phosphate [1,2,3,4]
The amino acid sequences of Teth514_1788 and Teth514_1789 showed no predicted Nterminal signal peptides based on a PSORTb Version 3.0.2 [18] and a SignalP 4.1 analysis [19]. These results suggest that Teth514_1788 and Teth514_1789 play a role in the intercellular phosphorolysis of certain b-mannoside
We identified Teth514_1788 and Teth514_1789 as new members of the GH130 phosphorylase, which catalyze the reversible phosphorolysis of 1,2-boligomannan
Summary
Glycoside phosphorylases catalyze the cleavage of glycosyl linkages via a substitution with inorganic phosphate [1,2,3,4]. These enzymes phosphorolyze particular glycosides to form corresponding sugar 1-phosphates with retention or PLOS ONE | DOI:10.1371/journal.pone.0114882. Because the phosphorylase reactions are reversible, various oligosaccharides have been synthesized via reverse phosphorolysis using the corresponding sugar 1-phosphate as a donor substrate and suitable carbohydrate acceptors [3, 4]. The discovery of a novel phosphorylase showing unreported substrate specificity and regioselectivity is desired to expand the number of synthesizable oligosaccharides
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