Abstract
Agarases catalyze the hydrolysis of agarose to oligosaccharides which display an array of biological and physiological functions with important industrial applications in health-related fields. In this study, the gene encoding agarase (Aga-ms-R) was cloned from Microbulbifer sp. BN3 strain. Sequence alignment indicated that Aga-ms-R belongs to the GH16 family and contains one active domain and two carbohydrate binding module (CBM) domains. The mature Aga-ms-R was expressed successfully by employing the Brevibacillus system. Purified rAga-ms-R was obtained with a specific activity of 100.75 U/mg. rAga-ms-R showed optimal activity at 50 °C and pH 7.0, and the enzyme activity was stable at 50 °C and also over the pH range of 5.0–9.0. After exposure of rAga-ms-R to 70 °C for 30 min, only partial enzyme activity remained. Thin layer chromatographic analysis of the enzymatic hydrolysate of agar obtained using rAga-ms-R disclosed that the hydrolysate comprised, in a long intermediate-stage of the hydrolysis reaction, mainly neoagarotetraose (NA4) and neoagarohexaose (NA6) but ultimately, predominantly neoagarotetraose and trace amounts of neoagarobiose (NA2). Hydrolysates of the raw red seaweeds Gracilaria sjoestedtii and Gelidium amansii, produced by incubation with rAga-ms-R, were mainly composed of neoagarotetraose. The results demonstrate the high efficiency of rAga-ms-R in producing neoagaraoligosaccharide under low-cost conditions.
Highlights
Agarase is a glycoside hydrolase (GH) that degrades agarose, and the bulk of the hydrolysate of agarose is composed of agarose oligosaccharide [1]
2.1. β-Agarase Gene Derived from Microbulbifer sp
Amino acid sequence alignment (Figure 1a) of Aga-ms-R indicates that it belongs to the GH16 family of β-agarases, with up to 97% similarity to β-agarase derived from Microbulbifer pacificus (WP_105101548.1, AYV64444.1); 56% similarity to β-agarase derived from Microbulbifer sp
Summary
Agarase is a glycoside hydrolase (GH) that degrades agarose, and the bulk of the hydrolysate of agarose is composed of agarose oligosaccharide [1]. Compared with methods of acidic hydrolysis, agarases catalyze the hydrolysis of agars/agarose to produce biologically improved oligosaccharides with notable efficiency and specificity, controllable products, simple production facilities, low energy consumption, and minimal environmental pollution are the advantages. The degradation products of agarose-catalyzed agarose hydrolysis in particular, display multiple biological and physiological activities including antioxidant and anti-inflammatory activities, with significant industrial applications in the cosmetic, pharmaceutical and food arenas [5,6]. Due to these favorable features of agar oligosaccharides, their preparation and potential applications have attracted widespread attention
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