INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
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2024
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IF 7.7
Wenhui Li,Yuqiu Wu,Si Long,Zhi Chen,Liangzhi Li,Xin Ju
The cascade of sugar isomerases is one of the most practical methods for producing rare sugars, and enzyme immobilization endows it with high economic efficiency, operational convenience and reusability. However, the most employed cross-linker glutaraldehyde (GA) has the disadvantages of enzyme deactivation and limitation of substrate binding. Herein, three compounds, glyoxal, GA, and 2,5-furandicarboxaldehyde (DFF) were evaluated within a previously developed cascade comprising ribose-5-phosphate isomerase and D-tagatose-3-epimerase to prepare D-ribulose form D-xylose. Analyses of surface morphology, element and chemical bond revealed that all compounds effectively cross-linked the isomerases. High concentration of the cross-linkers was generally beneficial for binding protein and preventing enzyme leak during reusing cycles. Glyoxal performed the highest immobilization rate, though it hadn't been employed as a cross-linker for enzyme immobilization. DFF mediated cross-linking revealed the highest activity recovery, substrate conversion and residual activity after reusing cycles, suggesting better biocompatibility than glyoxal and GA. After 8 rounds of recycling, the residual activity of enzyme immobilized by DFF was 61.4 %, ∼30 % higher than that of GA. This study proved a potential alternative cross-linker DFF for the immobilization of enzyme cascade with high activity recovery and reusability, which could promote the efficient production of high value-added products from biomass monosaccharides.
研究领域:
生物化学
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酶工程
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产品关联
DOI
PubMed
高影响因子
Molecular Catalysis
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2025
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IF 4.9
Si Long,Zhi Chen,Xin Ju,Liangzhi Li
Isomerization cascade is a promising strategy to prepare valuable rare sugars from biomass xylose. However current conversions are not satisfactory due to factors of enzyme efficiency and reaction equilibrium. An isomerase/isomerase cascade has been newly developed employing an isomerase of xylose and a tagatose-3-epimerase (DTE) to prepare xylulose and ribulose from xylose, with a total xylose conversion of 43.2 %. In this study, two microbial glucose isomerases (GI) were tested as alternatives to isomerize xylose for their high activity towards xylose and thermal stability, aiming to explore the adaption effect of enzyme in the isomerization type of cascade. GIs from Streptomyces rubiginosus (SrGI) and Piromyces sp. (PsGI) both showed higher xylose conversions than the previously used isomerase of xylose, ribose-5-phosphate isomerase. The optimal reaction conditions of GIs for xylose conversion were compared in the single transformation of xylose and the cascade reactions catalyzed by GI/DTE, and it revealed the importance of high reaction temperature to substrate conversion and enzyme deactivation. The activity profile and thermal stability of SrGI were better than PsGI, making it a potential biocatalyst in biomass sugar isomerization which was favorable at high temperatures. The conversion of xylose by the PsGI/DTE cascade was improved to 71.3 % by efforts of enzyme adaption and reaction condition adjusting. Additionally, the standstill of isomerization reaction of monosaccharide can be attributed to thermal dynamic equilibrium at low temperatures and enzyme deactivation at high temperatures based on the results of this study.
研究领域:
生物化学
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酶工程
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产品关联
DOI
高影响因子
ENZYME AND MICROBIAL TECHNOLOGY
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2021
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IF 3.493
Rong Wang,Xinqi Xu,Xuemei Yao,Hengtao Tang,Xin Ju,Liangzhi Li
Ribose-5-phosphate isomerase A (RpiA) is of great importance in biochemistry research, however its application in biotechnology has not been fully explored. In this study the activity of RpiA from Ochrobactrum sp. CSL1 (OsRpiA) towards D-allose was engineered based on sequential and structural analyses. Strategies of alanine scanning, rational design and saturated mutagenesis were employed to create three mutant libraries. A single mutant of K124A showed a 45 % activity improvement towards D-allose. The reaction properties of the mutant were analyzed, and a shift of optimal pH and higher thermal stability at low reaction temperatures were identified. The conversion of D-allose was also improved by 40 % using K124A, and higher activities on major substrates were found in the mutant’s substrate scope, implying its application potential in rare sugar preparation. Kinetics analysis revealed that K m of K124A mutant decreased by 12 % and the catalytic efficiency increased by 65 % towards D-allose. Moreover, molecular dynamics simulation illustrated the binding of substrate and K124A was more stable than that of the wild-type. The shorter distance and more relax bond angle between the catalytic residue of K124A and D-allose explained the activity improvement in detail. This study highlights the potential of OsRpiA as a biocatalyst for rare sugar preparation, and provides distinct evidences for its catalytic mechanism.
研究领域:
生物化学
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生物技术
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产品关联
DOI
PubMed
高影响因子
FOOD SCIENCE AND BIOTECHNOLOGY
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2023
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IF 2.9
Zheng Qian,Long Si,Chen Zhi,Fu Jiaolong,Ju Xin,Li Liangzhi
Enzymatic preparation of rare sugars as an alternative to traditional sweeteners is an effective strategy to achieve a low-calorie healthy diet. Ribose-5-phosphate isomerase B (RpiB) is a key enzyme in the non-oxidative branch of the catalytic pentose phosphate pathway. Here, we investigated the potential of Curtobacterium flaccumfaciens ZXL1 ( C. flaccumfaciens ZXL1) derived RpiB (CfRpiB) in D-allose preparation. The optimal reaction conditions for recombinant CfRpiB were found experimentally to be pH 7.0, 55 °C, and no metal ions. The kinetic parameters K m , k cat , and catalytic efficiency k ca t / K m were 320 mM, 4769 s −1 , and 14.9 mM −1 s −1 respectively. The conversion of D-allulose by purified enzyme (1 g L −1 ) to D-allose was 13% within 1 h. In addition, homology modeling and molecular docking were used to predict the active site residues: Asp13, Asp14, Cys72, Gly73, Thr74, Gly77, Asn106, and Lys144.
研究领域:
生物化学
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生物技术
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食品科学
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产品关联
DOI
PubMed
高影响因子
BIOORGANIC CHEMISTRY
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2024
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IF 4.5
Muhammad Waheed Iqbal,Xinrui Tang,Tahreem Riaz,Shahid Mahmood,Yufei Zhang,Mei Zhao,Junhua Yun,Jia Li,Xianghui Qi
6-Deoxy- l -sorbose (6-DLS) is an imperative rare sugar employed in food, agriculture, pharmaceutical and cosmetic industeries. However, it is a synthetic and very expensive rare sugars, previously synthesized by chemo-enzymatic methods through a long chain of chemical processes. Recently, enzymatic synthesis of rare sugars has attracted a lot of attention due to its advantages over synthetic methods. In this work, a promising approach for the synthesis of 6-DLS from an inexpensive sugar l -fucose was identified. The genes for l -fucose isomerase from Paenibacillus rhizosphaerae (Pr-LFI) and genes for d -tagatose-3-epimerase from Caballeronia fortuita (Cf-DTE) have been used for cloning and co-expression in Escherichia coli , developed a recombinant plasmid harboring pANY1-Pr-LFI/Cf-DTE vector. The recombinant co-expression system exhibited an optimum activity at 50 °C of temperature and pH 6.5 in the presence of Co 2+ metal ion which inflated the catalytic activity by 6.8 folds as compared to control group with no metal ions. The recombinant co-expressed system was stable up to more than 50 % relative activity after 12 h and revealed a melting temperature (T m ) of 63.38 °C exhibiting half-life of 13.17 h at 50 °C. The co-expression system exhibited, 4.93, 11.41 and 16.21 g/L of 6-DLS production from initial l -fucose concentration of 30, 70 and 100 g/L, which equates to conversion yield of 16.44 %, 16.30 % and 16.21 % respectively. Generally, this study offers a promising strategy for the biological production of 6-DLS from an inexpensive substrate l -fucose in slightly acidic conditions with the aid of co-expression system harboring Pr-LFI and CF-DTE genes.
研究领域:
生物技术
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酶学
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生化工程
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食品科学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
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