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Abstract

A DNA fragment, which complemented the growth of E. coli both on M9 medium containing raffinose and on LB medium containing ampicillin, IPTG and 5-bromo-4-chloro-3-indoxyl-alpha-D-galactoside, was isolated from the genomic library of Bifidobacterium longum SJ32, which had been digested with EcoRI. In the cloned DNA fragment, a gene encoding a sucrose phosphorylase (splP) and a partially cloned putative sucrose regulator gene (splR) were identified using the deletion analysis and sequence analysis. A 56 kDa protein was synthesized in E. coli and partially purified by DEAE-ion exchange chromatography. The partially purified enzyme did not react with melibiose, melezitoze and raffinose but did with sucrose. It had transglucosylation activity in addition to hydrolytic activity.

High-Yield Biosynthesis of Glucosylglycerol through Coupling Phosphorolysis and Transglycosylation Reactions

“…To found other GGPs with high activity, we also used the amino acid sequence of MaGGP as model and searched the NCBI database for candidate GGPs. Three possible interactions (Tyr194, Gln336, and Ala333) between the acceptor site residues and the glycerol moiety of glucosylglycerol have been identified according to the homology modelling and molecular docking between MaGGP and glucosylglycerol . We performed sequence alignment and three other enzymes with different sequence similarities, in which the amino acid residues around those three sites were conserved (Figure S1b), selected, and expressed in E.…”

“…Another enzyme sucrose phosphorylase (SP) from Leuconostoc mesenteroides , can transfer the glucosyl moiety from sucrose into the 2- O -position of glycerol, forming enantiomerically pure αGG. However, its transglycosylation activity is 4 orders of magnitude lower than that for sucrose phosphorolysis to α- d -glucose 1-phosphate (α-G1P), resulting in long reaction time for αGG synthesis. Recently, two glucosylglycerol phosphorylases (GGPs) from Bacillus selenitireducens , and Marinobacter adhaerens have been characterized with high catalytic ability to form αGG using β- d -glucose 1-phosphate (β-G1P) and α-G1P as sugar donors, respectively.…”

Sucrose 6F-phosphate phosphorylase: a novel insight in the human gut microbiome

“…(BAA14344.1 [17], ABS59292.1 [21], AAX33736.1 [23]), Streptococcus mutans (CAA30846.1 [16]), Bifidobacterium adolescentis (AAO33821.1 [20]), Bifidobacterium longum spp. (AAO84039.1 [19], BAF62433.1 [22]), Lactobacillus reuteri (AGK37834.1 [24]), Pelomonas saccharophila (AAD40317.1 [18]); 34 % to Tt SPP from Thermoanaerobacterium thermosaccharolyticum (ADL69407.1 [25]); 27 and 28 % to Ms GGaP and Ma GGoP (ADH62582.1 and ADP98617.1 [26, 27]), respectively (Fig. S5).…”

Genomic analysis of three Bifidobacterium species isolated from the calf gastrointestinal tract

“…The three calf strains ferment fructo-oligosaccharides (FOS, Table 2 ) and encode a three gene operon similar to that shown to be involved in fructo-oligosaccharide breakdown in B. breve and B. longum 42 . A sucrose utilization operon containing a GH13 sucrose phosphorylase/transglycosylase has been characterized in B. longum 43 and is present in AGR2137 and AGR2158. However, AGR2145 is unable to use sucrose ( Table 2 ) and the operon in this strain (G627DRAFT_00811-00815) contains an inserted toxin-antitoxin gene cassette ( Table S2 ).…”