The characterisation of an alkali-stable maltogenic amylase from Bacillus lehensis G1 and improved malto-oligosaccharide production by hydrolysis suppression.

Nor Hasmaliana Abdul Manas,Samson Pachelles,Nor Muhammad Mahadi,Rosli Md Illias,Alberto G. Passi

doi:10.1371/journal.pone.0106481

Nor Hasmaliana Abdul Manas, Samson Pachelles + Show 3 more

Open Access

https://doi.org/10.1371/journal.pone.0106481

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Abstract

A maltogenic amylase (MAG1) from alkaliphilic Bacillus lehensis G1 was cloned, expressed in Escherichia coli, purified and characterised for its hydrolysis and transglycosylation properties. The enzyme exhibited high stability at pH values from 7.0 to 10.0. The hydrolysis of β-cyclodextrin (β-CD) produced malto-oligosaccharides of various lengths. In addition to hydrolysis, MAG1 also demonstrated transglycosylation activity for the synthesis of longer malto-oligosaccharides. The thermodynamic equilibrium of the multiple reactions was shifted towards synthesis when the reaction conditions were optimised and the water activity was suppressed, which resulted in a yield of 38% transglycosylation products consisting of malto-oligosaccharides of various lengths. Thin layer chromatography and high-performance liquid chromatography analyses revealed the presence of malto-oligosaccharides with a higher degree of polymerisation than maltoheptaose, which has never been reported for other maltogenic amylases. The addition of organic solvents into the reaction further suppressed the water activity. The increase in the transglycosylation-to-hydrolysis ratio from 1.29 to 2.15 and the increased specificity toward maltopentaose production demonstrated the enhanced synthetic property of the enzyme. The high transglycosylation activity of maltogenic amylase offers a great advantage for synthesising malto-oligosaccharides and rare carbohydrates.

Highlights

Oligosaccharides offer numerous health benefits and improve the physicochemical properties of foods [1]
Maltogenic amylase, pullulanase (EC 3.2.1.4), cyclodextrin glucanotransferase (EC 2.4.1.19) and cyclodextrinase (EC 3.2.1.54) are known to exhibit different substrate specificities compared with a-amylase (EC 2.3.1.1), even though they are from the same family
Maltogenic amylase (MAG1) from B. lehensis G1 The gene sequence that corresponded to MAG1 consisted of

Summary

Introduction

Oligosaccharides offer numerous health benefits and improve the physicochemical properties of foods [1]. Maltogenic amylase (EC 3.2.1.133) is a potential catalyst for malto-oligosaccharide production It is an amylolytic enzyme from glycosyl hydrolase family 13 (GH13). Maltogenic amylase has been demonstrated to have a transglycosylation activity that forms sugar molecules of various lengths [5] This enzyme reportedly catalysed the formation of glycosidic linkages to produce oligosaccharides and various modified sugars [6,7,8]. No study on the influence of organic solvents on malto-oligosaccharide synthesis by maltogenic amylase has been reported to date. The products of b-CD hydrolysis and malto-oligosaccharide synthesis by transglycosylation were observed. This report is the first to describe the optimisation of reaction conditions and the incorporation of a water-miscible organic solvent to suppress hydrolysis activity during malto-oligosaccharide production by maltogenic amylase. The results presented here suggest that MAG1 is a promising candidate to catalyse the production of malto-oligosaccharides of various lengths

Materials and Methods

Results and Discussion

26.9 Tri- and tetra-saccharides