Levansucrases of a Pseudomonas syringae pathovar as catalysts for the synthesis of potentially prebiotic oligo- and polysaccharides

Visnapuu, Triinu; Mardo, Karin; Alamäe, Tiina

doi:10.1016/j.nbt.2015.01.009

Cited by 40 publications

(32 citation statements)

References 100 publications

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“…tomato and its mutant Asp300Asn [40] were used in this work to produce levans as substrates for the endo-levanase BT1760 (see Fig 1). The Lsc3 has a very high catalytic activity–the k cat for sucrose-splitting is over 500 1/s and the yield of levan in a 20-h reaction with 1.2 M sucrose is up to 13 g per 1 mg of Lsc3 protein [22]. Therefore, it is certainly feasible for biotechnological applications.…”

Section: Resultsmentioning

confidence: 99%

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A Highly Active Endo-Levanase BT1760 of a Dominant Mammalian Gut Commensal Bacteroides thetaiotaomicron Cleaves Not Only Various Bacterial Levans, but Also Levan of Timothy Grass

et al. 2017

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Bacteroides thetaiotaomicron, an abundant commensal of the human gut, degrades numerous complex carbohydrates. Recently, it was reported to grow on a β-2,6-linked polyfructan levan produced by Zymomonas mobilis degrading the polymer into fructooligosaccharides (FOS) with a cell surface bound endo-levanase BT1760. The FOS are consumed by B. thetaiotaomicron, but also by other gut bacteria, including health-promoting bifidobacteria and lactobacilli. Here we characterize biochemical properties of BT1760, including the activity of BT1760 on six bacterial levans synthesized by the levansucrase Lsc3 of Pseudomonas syringae pv. tomato, its mutant Asp300Asn, levansucrases of Zymomonas mobilis, Erwinia herbicola, Halomonas smyrnensis as well as on levan isolated from timothy grass. For the first time a plant levan is shown as a perfect substrate for an endo-fructanase of a human gut bacterium. BT1760 degraded levans to FOS with degree of polymerization from 2 to 13. At optimal reaction conditions up to 1 g of FOS were produced per 1 mg of BT1760 protein. Low molecular weight (<60 kDa) levans, including timothy grass levan and levan synthesized from sucrose by the Lsc3Asp300Asn, were degraded most rapidly whilst levan produced by Lsc3 from raffinose least rapidly. BT1760 catalyzed finely at human body temperature (37°C) and in moderately acidic environment (pH 5–6) that is typical for the gut lumen. According to differential scanning fluorimetry, the Tm of the endo-levanase was 51.5°C. All tested levans were sufficiently stable in acidic conditions (pH 2.0) simulating the gastric environment. Therefore, levans of both bacterial and plant origin may serve as a prebiotic fiber for B. thetaiotaomicron and contribute to short-chain fatty acids synthesis by gut microbiota. In the genome of Bacteroides xylanisolvens of human origin a putative levan degradation locus was disclosed.

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Section: Resultsmentioning

confidence: 99%

“…Levan-type fructans are mostly synthesized by bacterial enzymes [22], but are present also in some plants. For example, timothy grass ( Phleum pratense ) and orchard grass ( Dactylis glomerata ) have linear β-2,6-linked fructans which are referred to as phleins or plant levans [23–25].…”

Section: Introductionmentioning

confidence: 99%

A Highly Active Endo-Levanase BT1760 of a Dominant Mammalian Gut Commensal Bacteroides thetaiotaomicron Cleaves Not Only Various Bacterial Levans, but Also Levan of Timothy Grass

et al. 2017

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“…6A) cross-feeding of probiotic bacteria with levan degradation products (short-chain FOS) as 579 suggested in our previous studies (Adamberg et al, 2014;Visnapuu et al, 2015). On the basis 580 of these data, we hypothesize that B. thetaiotaomicron may enhance the bioavailability of 581 microelements (Co) and provide levan degradation products for the other members of gut 582 microbiota, whereas bacteria that are able to synthesize vitamin B 12 from Co, supply B.…”

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confidence: 70%

“…67 However, less information concerning functions and applications is available for levan 68 produced by various bacterial species. Inulin and levan are both fructans that differ according 69 to linkage type between the fructose residues in the main chain: β-2,1 in inulin and β-2,6 in 70 levan (Han, 1990;Visnapuu, Mardo, & Alamäe, 2015). Similarly to inulin, levan can be 71 potentially applied in medicine and food technology due to its prebiotic effect on beneficial 72 human gut bacteria ( levan-stabilized NPs with various physico-chemical methods and hypothesized that levan is 81…”

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Bacterial polysaccharide levan as stabilizing, non-toxic and functional coating material for microelement-nanoparticles

Bondarenko

Ivask

Kahru

et al. 2016

Carbohydrate Polymers

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“…Nevertheless, levansucrases of different origins have considerably different biochemical properties and product spectrum. Levansucrases of Gram-positive bacteria (such as B. subtilis, Bacillus megaterium and Streptococcus salivarius) produce generally levan without accumulating the intermediate FOS [14,15,16], while levansucrases from some Gram-negative bacteria (such as G. diazotrophicus, E. amylovora and Pseudomonas syringae) produce high amounts of FOS with small amounts of levan [17,18,19]. Structural studies recently pointed out that the main differences between FOS and levan-producing enzymes are located on the surface loops [12,20].…”

Section: Introductionmentioning

confidence: 99%

Structural Modelling and Structure-Function Analysis of <i>Zymomonas mobilis</i> Levansucrase

Bakar¹,

Türköz²

2017

SDÜ Fen Bil Enst Der

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Levansucrases are bacterial enzymes which produce fructan polymers from sucrose via hydrolysis and transfructosylation activities. These polymers; levan and fructooligosaccharides are valuable for food and pharmaceutical industries. Levansucrases from Gram-positive bacteria such as Bacillus subtilis tend to produce levan, while those from Gram-negative bacteria preferentially produce fructooligosaccharides. Zymomonas mobilis is an efficient levansucrase producer and its extracellular levansucrase can produce both fructooligosaccharides and levan depending on the reaction parameters. In this study, the structure of Z. mobilis levansucrase was modeled in order to help to understand the structure-function relationship of the enzyme. Furthermore, amino acids previously reported to be important for levansucrase activity were mapped on the model. The structural model presents a five-bladed propeller with a deep, negatively charged central pocket, similar to other bacterial levansucrases. Mapping showed that amino acids which previously reported to affect fructan length are located on the periphery of the structure covering the active site central pocket. Thus it is showed that, for the first time, that hydrolysis and transfructosylation reactions are catalyzed on different parts of Z. mobilis levansucrase structure. The structural location of the critical amino acids will pave the way to identify other residues which control fructan length by site directed mutagenesis without altering the overall fold of the enzyme.

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Levansucrases of a Pseudomonas syringae pathovar as catalysts for the synthesis of potentially prebiotic oligo- and polysaccharides

Cited by 40 publications

References 100 publications

A Highly Active Endo-Levanase BT1760 of a Dominant Mammalian Gut Commensal Bacteroides thetaiotaomicron Cleaves Not Only Various Bacterial Levans, but Also Levan of Timothy Grass

A Highly Active Endo-Levanase BT1760 of a Dominant Mammalian Gut Commensal Bacteroides thetaiotaomicron Cleaves Not Only Various Bacterial Levans, but Also Levan of Timothy Grass

Bacterial polysaccharide levan as stabilizing, non-toxic and functional coating material for microelement-nanoparticles

Structural Modelling and Structure-Function Analysis of <i>Zymomonas mobilis</i> Levansucrase

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