Α-Galactobiosyl units: thermodynamics and kinetics of their formation by transglycosylations catalysed by the GH36 α-galactosidase from Thermotoga maritima. (12th January 2015)

• Record Type: Journal Article
• Authors: Borisova, Anna S.; Ivanen, Dina R.; Bobrov, Kirill S.; Eneyskaya, Elena V.; Rychkov, Georgy N.; Sandgren, Mats; Kulminskaya, Anna A.; Sinnott, Michael L.; Shabalin, Konstantin A.
• Is Part Of: Carbohydrate research. Volume 401(2015)
• Page Start: 115
• Page End: 121
• Publication Date: 2015-01-12
• Subjects: TmGal36A α galactosidase from Thermotoga maritima -- pNP para-nitrophenol -- pNPGal pNP-α-d-galactopyranoside -- 1, 2diGal α-d-Galp-(1 → 2)-α-d-Galp -- 1, 3diGal α-d-Galp-(1 → 3)-α-d-Galp -- 1, 4diGal α-d-Galp-(1 → 4)-α-d-Galp -- 1, 6diGal α-d-Galp-(1 → 6)-α-d-Galp -- pNP1, 2diGal α-d-Galp-(1 → 2)-α-d-Galp-O-pNP -- pNP1, 3diGal α-d-Galp-(1 → 3)-α-d-Galp-O-pNP -- pNP1, 6diGal α-d-Galp-(1 → 6)-α-d-Galp-O-pNP -- pNPdiGal pNP-digalactopyranoside; α-Galactosidase -- Thermotoga maritima -- Regioselectivity -- Transglycosylation -- Kinetics -- Free energy differences; Carbohydrates -- Periodicals; Chemistry, Organic -- Periodicals; Biochemistry -- Periodicals; Carbohydrates -- Periodicals; Chimie organique -- Périodiques; Glucides -- Périodiques; Biochemistry; Carbohydrates; Chemistry, Organic; Periodicals; Electronic journals; 507.78
• DOI: 10.1016/j.carres.2014.11.003
• Languages: English
• ISSNs: 0008-6215

Abstract

Graphical abstract: Highlights: Total curves of transglycosylation yield kinetic parameters in a single experiment. Thermodynamics and relative stability of a glycosidic linkage are given. The synthesis of α1, 2-, α1, 3-, α1, 4- and α1, 6-galactobiosides is reported. Abstract: Broad regioselectivity of α-galactosidase from Thermotoga maritima ( Tm Gal36A) is a limiting factor for application of the enzyme in the directed synthesis of oligogalactosides. However, this property can be used as a convenient tool in studies of thermodynamics of a glycosidic bond. Here, a novel approach to energy difference estimation is suggested. Both transglycosylation and hydrolysis of three types of galactosidic linkages were investigated using total kinetics of formation and hydrolysis of p NP-galactobiosides catalysed by monomeric glycoside hydrolase family 36 α-galactosidase from T. maritima, a retaining exo -acting glycoside hydrolase. We have estimated transition state free energy differences between the 1, 2- and 1, 3-linkage (ΔΔ G ‡ 0 values were equal 5.34 ± 0.85 kJ/mol) and between 1, 6-linkage and 1, 3-linkage (ΔΔ G ‡ 0 = 1.46 ± 0.23 kJ/mol) in p NP-galactobiosides over the course of the reaction catalysed by Tm Gal36A. Using the free energy difference for formation and hydrolysis of glycosidic linkages (ΔΔ G ‡ F − ΔΔ G ‡ H ), we found that the 1, 2-linkage was 2.93 ± 0.47 kJ/mol higher in free energy than the 1, 3-linkage, and the 1, 6-linkage 4.44 ± 0.71 kJ/mol lower.