Highly Hydrolytic Reuteransucrase from Probiotic Lactobacillus reuteri Strain ATCC 55730

Abstract

Lactobacillus reuteri strain ATCC 55730 (LB BIO) was isolated as a pure culture from a Reuteri tablet purchased from the BioGaia company. This probiotic strain produces a soluble glucan (reuteran), in which the majority of the linkages are of the ␣-(134) glucosidic type (ϳ70%). This reuteran also contains ␣-(136)-linked glucosyl units and 4,6-disubstituted ␣-glucosyl units at the branching points. The LB BIO glucansucrase gene (gtfO) was cloned and expressed in Escherichia coli, and the GTFO enzyme was purified. The recombinant GTFO enzyme and the LB BIO culture supernatants synthesized identical glucan polymers with respect to linkage type and size distribution. GTFO thus is a reuteransucrase, responsible for synthesis of this reuteran polymer in LB BIO. The preference of GTFO for synthesizing ␣-(134) linkages is also evident from the oligosaccharides produced from sucrose with different acceptor substrates, e.g., isopanose from isomaltose. GTFO has a relatively high hydrolysis/transferase activity ratio. Complete conversion of 100 mM sucrose by GTFO nevertheless yielded large amounts of reuteran, although more than 50% of sucrose was converted into glucose. This is only the second example of the isolation and characterization of a reuteransucrase and its reuteran product, both found in different L. reuteri strains. GTFO synthesizes a reuteran with the highest amount of ␣-(134) linkages reported to date.

Potentials of Exopolysaccharides from Lactic Acid Bacteria

“…It has 70% a-1,4 linkage, also a-1,6 glycosidic bonds and 16% 4,6-disubstituted a-glucosyl units at the branching points and molecular weight of 40 MDa [27]. It is elaborated by Lactobacillus reuteri strain LB 121, Lactobacillus reuteri strain ATCC 55730 and Lactobacillus reuteri strain 35-5 have been reported to produce reuteran.…”

Structure-Function Relationships of Glucansucrase and Fructansucrase Enzymes from Lactic Acid Bacteria

“…Koepsell et al (89) observed that in the presence of sucrose and saccharide acceptor substrates such as maltose, isomaltose, and O-␣-methylglucoside, GS enzymes shift from glucan synthesis towards the synthesis of oligosaccharides (the acceptor reaction). Most acceptor reaction studies have been performed using saccharides (5,42,43,53,94,99) or saccharide derivatives as substrates (31,36,150). However, aromatic compounds (e.g., catechine) and salicyl alcohol have also been shown to act as acceptor substrates (114,221).…”

“…In order to elucidate which enzyme activities are encoded, these genes were cloned and heterologously expressed. The distribution of glucosidic linkages has been elucidated for the glucans synthesized from sucrose by heterologously produced (mostly using Escherichia coli as the host) GS enzymes from (i) seven Streptococcus strains (13 GS enzymes, synthesizing either dextran or mutan polymers) (67,90,124), (ii) four Leuconostoc strains (seven GS enzymes, synthesizing mainly dextran polymers, but also one alternan, and a GS synthesizing large numbers of ␣-(132,6) branch points) (4,17,55,124,135), and (iii) seven Lactobacillus strains (seven GS enzymes, synthesizing mainly dextrans, but also reuteran, and a highly branched mutan) (94,95,97) (Table 1).…”

Binding Interactions Between α-glucans from Lactobacillus reuteri and Milk Proteins Characterised by Surface Plasmon Resonance

“…Production and isolation of the α-glucans have previously been described. 7,22,23 Milk Proteins β-lactoglobulin and κ-casein were purchased from Sigma-Aldrich (St. Louis, MO, USA). β-lactoglobulin (5 gL -1 ) was heat-denatured (85ºC, 15 min) in sodium phosphate (50 mM, 30 mM NaCl, pH 6.8).…”

“…In the present study, glucans from different strains of Lactobacillus reuteri (ATCC-55730, 180, ML1 and 121) of varying molecular weight, linkage types, and degree of branching 7,[21][22][23][24] were analysed for binding to native β-lactoglobulin, denatured β-lactoglobulin and κ-casein in the range pH 4.0-5.5 by using SPR. The extent of individual interactions was correlated with the molecular properties of the glucans.…”