Rational re-design of <i>Lactobacillus reuteri</i> 121 inulosucrase for product chain length control

Charoenwongpaiboon, Thanapon; Klaewkla, Methus; Chunsrivirot, Surasak; Wangpaiboon, Karan; Pichyangkura, Rath; Field, Robert A.; Prousoontorn, Manchumas Hengsakul

doi:10.1039/c9ra02137j

Cited by 24 publications

(7 citation statements)

References 29 publications

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“…Recently, we reported a series of mutations of L. reuteri 121 inulosucrase, with the mutant R483A (R483A-LrInu) showing potential application for the production of IFOs [26,27]. In the present study, R483A-LrInu was prepared as a CLEAs format, with the optimization of reaction conditions.…”

Section: Of 12mentioning

confidence: 94%

Preparation of Cross-Linked Enzyme Aggregates (CLEAs) of an Inulosucrase Mutant for the Enzymatic Synthesis of Inulin-Type Fructooligosaccharides

et al. 2019

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Fructooligosaccharides are well-known carbohydrate molecules that exhibit good probiotic activity and are widely used as sweeteners. Inulin-type fructooligosaccharides (IFOs) can be synthesized from sucrose using inulosucrase. In this study, cross-linked enzyme aggregates (CLEAs) of Lactobacillus reuteri 121 inulosucrase (R483A-LrInu) were prepared and used as a biocatalyst for IFOs production. Under optimum conditions, R483A-LrInu CLEAs retained 42% of original inulosucrase activity. Biochemical characterization demonstrated that the optimum pH of inulosucrase changed from 5 to 4 after immobilization, while the optimum temperature was unchanged. Furthermore, the pH stability and thermostability of the R483A-LrInu CLEAs was significantly improved. IFOs product characterization indicated that the product specificity of the enzyme was impacted by CLEA generation, producing a narrower range of IFOs than the soluble enzyme. In addition, the R483A-LrInu CLEAs showed operational stability in the batch synthesis of IFOs.

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Section: Of 12mentioning

confidence: 94%

Preparation of Cross-Linked Enzyme Aggregates (CLEAs) of an Inulosucrase Mutant for the Enzymatic Synthesis of Inulin-Type Fructooligosaccharides

et al. 2019

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“…LEaP module in AMBER18 with GLYCAM06j-1 force field and ff14SB parameters [ 58 ] was used to immerse the representative binding orientations of all systems in isomeric truncated octahedral boxes of TIP3P water molecules with the 13 Å buffer distance. The five-step minimization procedure [ 58 , 59 , 60 , 61 , 62 , 63 , 64 , 65 , 66 , 67 , 68 , 69 , 70 , 71 , 72 , 73 ] was employed to reduce the unfavorable interaction of the system. In this study, all steps used 2,500 steps of steepest descent and 2,500 steps of conjugate gradient.…”

Section: Methodsmentioning

confidence: 99%

Biochemical and ligand binding properties of recombinant Xenopus laevis cortical granule lectin-1

Deetanya¹,

Sitthiyotha²,

Chomanee³

et al. 2022

Heliyon

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“…The TLC analysis was performed according to the method described previously [24]. The mobile phase consisted of 1-butanol: glacial acetic acid: water, 3:3:2 (v/v/v).…”

Section: Thin-layer Chromatography (Tlc)mentioning

confidence: 99%

Levansucrase from Bacillus amyloliquefaciens KK9 and Its Y237S Variant Producing the High Bioactive Levan-Type Fructooligosaccharides

et al. 2020

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Levan-typed fructooligosaccharide (LFOS), a β-2,6 linked oligofructose, displays the potential application as a prebiotic and therapeutic dietary supplement. In the present study, LFOS was synthesized using levansucrase from Bacillus amyloliquefaciens KK9 (LsKK9). The wild-type LsKK9 was cloned and expressed in E. coli, and purified by cation exchanger chromatography. Additionally, Y237S variant of LsKK9 was constructed based on sequence alignment and structural analysis to enhance the LFOS production. High-performance anion-exchange chromatography coupled with pulsed amperometric detection (HPAEC-PAD) analysis indicated that Y237S variant efficiently produced a higher amount of short-chain LFOS than wild type. Also, the concentration of enzyme and sucrose in the reactions was optimized. Finally, prebiotic activity assay demonstrated that LFOS produced by Y237S variant had higher prebiotic activity than that of the wild-type enzyme, making the variant enzyme attractive for food biotechnology.

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Rational re-design of Lactobacillus reuteri 121 inulosucrase for product chain length control

Abstract: Blocking the binding track of inulosucrase to control the chain length of the products.

Cited by 24 publications

References 29 publications

Preparation of Cross-Linked Enzyme Aggregates (CLEAs) of an Inulosucrase Mutant for the Enzymatic Synthesis of Inulin-Type Fructooligosaccharides

Preparation of Cross-Linked Enzyme Aggregates (CLEAs) of an Inulosucrase Mutant for the Enzymatic Synthesis of Inulin-Type Fructooligosaccharides

Biochemical and ligand binding properties of recombinant Xenopus laevis cortical granule lectin-1

Levansucrase from Bacillus amyloliquefaciens KK9 and Its Y237S Variant Producing the High Bioactive Levan-Type Fructooligosaccharides

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