Mode of action of acetylxylan esterase from Streptomyces lividans: a study with deoxy and deoxy-fluoro analogues of acetylated methyl β- d-xylopyranoside

• Published in: Biochimica et biophysica acta Vol. 1622; no. 2; pp. 82 - 88
• Main Authors: Biely, Peter, Mastihubová, Mária, Côté, Gregory L, Greene, Richard V
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 23.07.2003
• Subjects: Acetates - chemistry; Acetylation; Acetylesterase - chemistry; Acetylesterase - metabolism; Acetylxylan esterase; Deacetylation of carbohydrate; enzyme inhibition; enzyme substrates; esterases; Methylglycosides - chemistry; Methylglycosides - metabolism; Molecular Conformation; nuclear magnetic resonance spectroscopy; Streptomyces - enzymology; Streptomyces lividans; Substrate Specificity; AcXE; 2-de-2-F-3,4-di- O-Ac-Me-β-Xyl p; Acetylxylan esterase; Me-β-Xyl p; Streptomyces lividans; 2,3,4-tri- O-Ac-Me-β-Xyl p; 2-de-3,4-di- O-Ac-Me-β-Xyl p; 2,4-di- O-Ac-Me-β-Xyl p; Deacetylation of carbohydrate
• ISSN: 0304-4165; 0006-3002; 1872-8006; 1878-2434
• DOI: 10.1016/S0304-4165(03)00130-2

Streptomyces lividans acetylxylan esterase removes the 2- or 3- O-acetyl groups from methyl 2,4-di- O-acetyl- and 3,4-di- O-acetyl β- d-xylopyranoside. When the free hydroxyl group was replaced with a hydrogen or fluorine, the rate of deacetylation was markedly reduced, but regioselectivity was not affected. The regioselectivity of deacetylation was found to be independent of the prevailing conformation of the substrates in solution as determined by 1H-NMR spectroscopy. These observations confirm the importance of the vicinal hydroxyl group and are consistent with our earlier hypothesis that the deacetylation of positions 2 and 3 may involve a common ortho-ester intermediate. Another possible role of the free vicinal hydroxyl group could be the activation of the acyl leaving group in the deacetylation mechanism. Involvement of the free hydroxyl group in the enzyme–substrate binding is not supported by the results of inhibition experiments in which methyl 2,4-di- O-acetyl β- d-xylopyranoside was used as substrate and its analogues or methyl β- d-xylopyranoside as inhibitors. The enzyme requires for its efficient action the trans arrangement of the free and acetylated hydroxyl groups at positions 2 and 3.