A Novel Agarolytic β-Galactosidase Acts on Agarooligosaccharides for Complete Hydrolysis of Agarose into Monomers

Abstract

cMarine red macroalgae have emerged to be renewable biomass for the production of chemicals and biofuels, because carbohydrates that form the major component of red macroalgae can be hydrolyzed into fermentable sugars. The main carbohydrate in red algae is agarose, and it is composed of D-galactose and 3,6-anhydro-L-galactose (AHG), which are alternately bonded by ␤1-4 and ␣1-3 linkages. In this study, a novel ␤-galactosidase that can act on agarooligosaccharides (AOSs) to release galactose was discovered in a marine bacterium (Vibrio sp. strain EJY3); the enzyme is annotated as Vibrio sp. EJY3 agarolytic ␤-galactosidase (VejABG). Unlike the lacZ-encoded ␤-galactosidase from Escherichia coli, VejABG does not hydrolyze common substrates like lactose and can act only on the galactose moiety at the nonreducing end of AOS. The optimum pH and temperature of VejABG on an agarotriose substrate were 7 and 35°C, respectively. Its catalytic efficiency with agarotriose was also similar to that with agaropentaose or agaroheptaose. Since agarotriose lingers as the unreacted residual oligomer in the currently available saccharification system using ␤-agarases and acid prehydrolysis, the agarotriose-hydrolyzing capability of this novel ␤-galactosidase offers an enormous advantage in the saccharification of agarose or agar in red macroalgae for its use as a biomass feedstock for fermentable sugar production.

Red macroalgae as a sustainable resource for bio-based products

“…The AOSs are enzymatically hydrolyzed to neoagarobiose and the monomeric sugars, AHG and D-galactose, by the sequential use of b-agarase II (e.g., Aga50D) and a-neoagarobiose hydrolase (NABH) [3,4]. A drawback of this process is that agarotriose (galactosyl-b-1,4-3,6-anhydro-L-galactosyl-a-1,3-galactose), the smallest odd-numbered AOS which is produced by b-agarase II, is not further hydrolyzed by typical b-agarases and NABH [3,5]. Recently, a novel agarolytic b-galactosidase (ABG) that acts on AOSs including agarotriose to release D-galactose was identified from a marine bacterium ( Figure 2B) [5].…”

“…A drawback of this process is that agarotriose (galactosyl-b-1,4-3,6-anhydro-L-galactosyl-a-1,3-galactose), the smallest odd-numbered AOS which is produced by b-agarase II, is not further hydrolyzed by typical b-agarases and NABH [3,5]. Recently, a novel agarolytic b-galactosidase (ABG) that acts on AOSs including agarotriose to release D-galactose was identified from a marine bacterium ( Figure 2B) [5]. The application of ABG to the saccharification of agarose increased the final yield of monomeric sugar, mainly due to the hydrolysis of agarotriose [5].…”

“…Recently, a novel agarolytic b-galactosidase (ABG) that acts on AOSs including agarotriose to release D-galactose was identified from a marine bacterium ( Figure 2B) [5]. The application of ABG to the saccharification of agarose increased the final yield of monomeric sugar, mainly due to the hydrolysis of agarotriose [5].…”

The novel catabolic pathway of 3,6‐anhydro‐L‐galactose, the main component of red macroalgae, in a marine bacterium

“… Saccharification and conversion of red macroalgae into fuels and chemicals. ( A ) Agarose, the major carbohydrate of red macroalgae, is depolymerized into agarooligosaccharides with main degrees of polymerization ( DP ) 4, 6, 8 and 10 by acid liquefaction using acetic acid ( L ee et al ., ), and these agarooligosaccharides are further enzymatically hydrolysed into AHG and galactose using β‐agarases and α‐neoagarobiose hydrolase (NABH). ( B ) The incorporation of the novel catabolic pathway of AHG into fermentative microorganisms enables the enhanced production of fuels and chemicals from red macroalgae.…”

A Novel Glycoside Hydrolase Family 5 β-1,3-1,6-Endoglucanase from Saccharophagus degradans 2-40 T and Its Transglycosylase Activity

“…The spot was rapidly dried under vacuum for homogeneous crystallization. The samples were analyzed by MALDI-TOF/TOF MS as described previously (27). Raw MS data were processed using FlexAnalysis software (version 3.3; Bruker Daltonics).…”