RSC Advances
·
2018
·Vol. 8
·期 72
·41247-41254
·
IF 2.936
Jiong-feng Zhao,Zhe Wang,Fei-long Gao,Jian-ping Lin,Li-rong Yang,Mian-bin Wu
Rhizopus oryzae lipase (ROL) is important because of its extreme sn-1,3-regioselectivity, but it shows poor thermostability, which severely restricts its application. In this work, the thermostability of ROL was greatly enhanced by rational design. First, several sites that may affect the thermostability of ROL were identified by multiple-sequence alignment. The half-lives of mutants V209L and D262G at 55 °C were about 4.38- and 4.2-times those of the wild-type, respectively. Then, a disulfide bond was introduced between positions 190 and 238 based on the prediction of Disulfide by Design 2, which greatly improved the thermostability of the protein. The activity of variant E190C/E238C retained about 58.2% after incubation at 55 °C for 720 min, whereas the half-life of wild type ROL was only about 11.7 min. On the basis of the results obtained by the two methods, we carried out a combined mutation. Quadruple mutant V209L/D262G/E190C/E238C was constructed and the thermostability was improved even further. The half-lives at 55 °C and 65 °C were 102.5- and 20-times those of the wild-type ROL. This improvement in thermostability will give ROL wider industrial applicability, especially in the preparation of structured lipids.
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BIOPROCESS AND BIOSYSTEMS ENGINEERING
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2014
·Vol. 37
·期 8
·1617-1626
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IF 1.823
Hu Yi,Yang Jiao,Jia Ru,Ding Yan,Li Shuang,Huang He
Chemical modification of lysine residues in Candida rugosa lipase (CRL) was carried out using five different functional ionic liquids, and about 15.4–25.0 % of the primary amino groups of lysine were modified. Enzymatic properties of the native and modified CRLs were investigated in olive oil hydrolysis reaction. Improved thermal stability, catalytic activity in organic solvents, and adaptability to temperature and pH changes were achieved compared with the native enzyme. CRL modified by [choline][H 2 PO 4 ] showed the best results, bearing a maximum improvement of 16.7 % in terms of relative activity, 5.2-fold increase in thermostability (after incubation at 45 °C for 5 h), and 2.3-fold increase in activity in strong polar organic solvent (80 % dimethyl sulfoxide) compared with the native enzyme. The results of ultraviolet, circular dichroism and fluorescence spectroscopy suggested that the change of the secondary and tertiary structures of CRL caused by the chemical modification resulted in the enhancement of enzymatic performance. The modification of CRL with functional ionic liquids was proved to be a novel and efficient method for improving the enzymatic properties of CRL.
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CHINESE JOURNAL OF CHEMICAL ENGINEERING
·
2014
·
IF 0.872
Jingyun Wang,Fangling Ji,Jishuang Xing,Shuang Cui,Yongming Bao,Wenbo Hao
A block copolymer of 2-dimethylaminoethyl methacrylate (DMAEMA) and glycidyl methacrylate (GMA) was grafted onto the surface of magnetic nanoparticles (Fe 3 O 4 ) via atom transfer radical polymerization . The resultant PGMA-b-PDMAEMA-grafted-Fe 3 O 4 magnetic nanoparticles with amino and epoxy groups were characterized by Fourier transform infrared spectroscopy, powder X-ray diffraction, thermo-gravimetric analysis, and scanning electron microscopy. Lipase from Burkholderia cepacia was successfully immobilized onto the magnetic nanoparticles by physical adsorption and covalent bonding. The immobilization capacity of the magnetic particles is 0.5 mg lipase per mg support, with an activity recovery of up to 43.1% under the optimum immobilization condition. Biochemical characterization shows that the immobilized lipase exhibits improved thermal stability, good tolerance to organic solvents with high lg P , and higher pH stability than the free lipase at pH 9.0. After six consecutive cycles, the residual activity of the immobilized lipase is still over 55% of its initial activity.
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RSC Advances
·
2015
·Vol. 5
·期 29
·23039-23045
·
IF 3.84
Weiwei Zhu,Yijing Li,Fang Zeng,Hang Yin,Liyuan Wang,Hao Zhu
Superparamagnetic Fe3O4 nanoparticles with diameter about 18 nm were prepared by a solvothermal method. A simple and green method was used to prepare amino-rich magnetic nanoparticles. The magnetic nanoparticles were modified by polyethylenimine (PEI) which is a polycationic polymer when pH < 10. The resulted magnetic nanoparticles were activated by glutaraldehyde to obtain nanoscale support for Candida rugosa Lipase (CRL) immobilization. The CRL was immobilized on the magnetic nanoparticles covalently, as well as via ionic exchange. The structure and magnetic behavior of the magnetic nanoparticles were confirmed by transmission electron microscopy (TEM), Fourier transform infrared spectroscopy (FT-IR), and vibrating sample magnetometer (VSM). Then the properties of the immobilized CRL were investigated, and the results showed that the obtained immobilized lipase displayed good reusability and applicability. The immobilized Candida rugosa Lipase (ICRL) presented wider pH tolerance (residual activity > 80% in a pH range from 5.5 to 8.0) than free Candida rugosa Lipase (residual activity decreased rapidly when the pH values were away from 7.0). ICRL showed excellent thermal stability (the relative of ICRL > 90%) after keeping in the water bath at 50 °C for 150 min while free Candida rugosa Lipase (FCRL) was absolutely deactivated. After being reused 10 times, ICRL maintained 60% relative activity.
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ORGANIC & BIOMOLECULAR CHEMISTRY
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2013
·Vol. 11
·期 41
·7192-7198
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IF 3.568
Ru Jia,Yi Hu,Luo Liu,Ling Jiang,He Huang
Various imidazolium-functional ionic liquids (ILs) composed of different cations and anions were grafted onto Candida antarctica lipase B (CALB) through lysine coupling, and 4–6 of the 9 primary amino groups of the lysine residues were modified. The catalytic activity and stability were investigated in a p-nitrophenyl palmitate hydrolysis reaction. After modification, CALB was activated and achieved a high catalytic efficiency in the aqueous phase (e.g., [HOOCMMIm][Cl] modification showed a 1.5-fold increase in the catalytic efficiency). The thermostability and organic solvent tolerance were significantly increased with the ILs modification (e.g., [HOOCBMIm][Cl] owned kosmotropic cation and chaotropic anion modification showed a 7-fold thermostability increase at 70 °C, 1-fold increase in 50% aqueous dimethylformamide and 5-fold increase in 50% aqueous methanol). Conformation changes were confirmed by fluorescence spectroscopy, circular dichroism spectroscopy and attenuated total reflection Fourier transform infrared spectroscopy.
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POLYMER
·
2015
·
IF 3.562
Wan-Xia Wu,Lin Qu,Bei-Yu Liu,Wei-Wei Zhang,Na Wang,Xiao-Qi Yu
A novel linear polyester with acid-labile β -thiopropionate groups was synthesized via the direct lipase-catalyzed polycondensation of methyl 3-((2-hydroxyethyl)thio)propanoate (MHETP) using immobilized lipase B from Candida antarctica (CALB). The influences of reaction conditions such as polymerization temperature and time on the molecular weight of the poly( β -thioether ester) (PTE) were studied. Ring-opening and condensation copolymerization of MHETP with ε -caprolactone (CL) led to the synthesis of novel poly( β -thioether ester- co - ε -caprolactone) copolymers , poly(TE- co -CL), with well-defined composition and narrow polydispersity . The random copolymers were characterized by 1 H NMR, 13 C NMR, 2D NMR , GPC , TGA and DSC . The degradation of the poly( β -thioether ester) under acidic conditions was investigated by GPC and 1 H NMR spectroscopy.
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JOURNAL OF NANOPARTICLE RESEARCH
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2017
·Vol. 19
·期 8
·1-11
·
IF 2.02
Cheong Ling-Zhi,Wei Yayu,Wang Hongbin,Wang Zhiying,Su Xiurong,Shen Cai
Zeolitic imidazolate frameworks (ZIF) represent one of the metal organic frameworks (MOF) with high potential for enzyme immobilization due to their exceptional chemical and thermal stability, negligible cytotoxicity, and easy synthesis under mild biocompatible conditions. Amine-functionalized ZIF-8 (An-ZIF-8) are capable of forming multipoint attachment via hydrogen bonding with lipase which will immobilize and further enhance stabilization of lipase. In addition, increased hydrophilicity of An-ZIF-8 will increase partitioning of An-ZIF-8 immobilized lipase at the aqueous/organic interface which enable lipase to expose its active site and retain its catalytic activity at its highest. Present study reports the use of ZIF-8 and An-ZIF-8 nanoparticles as carrier for Burkholderia cepacia lipase (BCL), compares the ester hydrolysis and transesterification activities of immobilized lipase with those of free lipase, and evaluates the reusability and recovery rate of the immobilized lipase. An-ZIF-8 nanoparticles (average 130.42 ± 0.55 nm) were facilely synthesized via mixing ZIF-8 nanoparticles with ammonia hydroxide solution. Despite having similar characteristics of high crystallinity and forming cuboid-like particles, An-ZIF-8 demonstrated significantly ( P < 0.05) lower Brunauer-Emmett-Teller (BET) surface area and higher thermal stability than ZIF-8. BCL were successfully immobilized on ZIF-8 (BCL@ZIF-8) and An-ZIF-8 (BCL@An-ZIF-8) nanoparticles with an average lipase loading rate of ~ 8 mg/g MOF. The immobilized BCL demonstrated no significant differences in terms of esters hydrolysis and transesterification activities with those of free BCL. BCL@An-ZIF-8 demonstrated superior catalytic stability in comparison to BCL@ZIF-8 with retainment of more than 80% of its initial hydrolysis and transesterification activity for at least 10 repeated runs. In addition, more than 80% of the BCL@An-ZIF-8 can be easily recovered during each cycle of the reusability test through simple centrifugation.
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BIOCHEMICAL ENGINEERING JOURNAL
·
2017
·
IF 2.892
Guanglei Zhao,Xiongfei Lang,Fengli Wang,Junrong Li,Xiaofeng Li
Long-chain fatty acid chitosan esters are important chitosan products with attractive application prospects in biological and biomedical applications. A new enzymatic method for synthesis of chitosan esters via transesterification with methyl palmitate has been developed. A reaction system containing a hydrophilic ionic liquid (IL) 1-ethyl-3-methyl imidazoliumacetate and a hydrophobic IL 1-Butyl-3-methylimidazolium tetrafluoroborate enables the transesterification of chitosan with high DS and selectivity in a homogeneous phase. FT-IR and NMR analysis confirmed that the chitosan ester was formed and the lipase selectively catalyzed the acylation of the 6-OH groups of chitosan without protection of its −NH 2 groups. The effects of several key reaction factors were investigated and the maximal degree of substitution (DS) value of 0.214 was obtained under the optimum conditions. The X-ray diffraction analysis showed that morphological and crystallographic properties of native chitosan were largely destroyed after modification. Thermogravimetric analysis showed that the thermal stability of chitosan palmitate was reduced. This is the first time that long-chain chitosan esters was biocatalytically synthesized in ILs and the biocatalytic method presented here showed potential application for chitosan modification processes of industrial interest.
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CHINESE JOURNAL OF CATALYSIS
·
2016
·
IF 2.628
Dun Deng,Yun Zhang,Aijun Sun,Yunfeng Hu
We previously functionally characterized a novel marine microbial GDSL lipase MT6 and identified that the stereo-selectivity of MT6 was opposite to that of other common lipases in trans-esterification reactions. Herein, we have investigated the use of MT6 in stereo-selective biocatalysis through direct hydrolysis reactions. Notably, the stereo-selectivity of MT6 was also demonstrated to be opposite to that of other common lipases in hydrolysis reactions. Parameters, including temperature, organic co-solvents, pH, ionic strength, catalyst loading, substrate concentration, and reaction time, affecting the enzymatic resolution of racemic 1-phenylethyl acetate were further investigated, with the e.e. of the final ( S )-1-Phenylethanol product and the conversion being 97% and 28.5%, respectively, after process optimization. The lengths of side chains of 1-phenylethyl esters greatly affected the stereo-selectivity and conversion during kinetic resolutions. MT6 is a novel marine microbial GDSL lipase exhibiting opposite stereo-selectivities than other common lipases in both trans-esterification reactions and hydrolysis reactions.
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FOOD CHEMISTRY
·
2016
·
IF 4.052
Xiumei Wang,Daoming Li,Man Qu,Rabia Durrani,Bo Yang,Yonghua Wang
Immobilization of lipase MAS1 from marine Streptomyces sp. strain W007 and its application in catalyzing esterification of n-3 polyunsaturated fatty acids (PUFA) with glycerol were investigated. The resin XAD1180 was selected as a suitable support for the immobilization of lipase MAS1, and its absorption ability was 75 mg/g (lipase/resin ratio) with initial buffer pH value of 8.0. The thermal stability of immobilized MAS1 was improved significantly compared with that of the free lipase. Immobilized MAS1 had no regiospecificity in the hydrolysis of triolein. The highest esterification degree (99.31%) and TAG content (92.26%) by immobilized MAS1-catalyzed esterification were achieved under the optimized conditions, which were significantly better than those (82.16% and 47.26%, respectively) by Novozym 435. More than 92% n-3 PUFA was incorporated into TAG that had similar fatty acids composition to the substrate (n-3 PUFA). The immobilized MAS1 exhibited 50% of its initial activity after being used for five cycles.
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INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
·
2025
·
IF 8.5
Hao Li,Bizhu Sun,Meiai Huang,Yipeng Liu,Qian Zhang,Zhuolin Luo,Quan Zeng,Wenjing Zhu,Xuan Li,Juan Chen,Xin Yuan,Panliang Zhang,Kewen Tang
The development of efficient immobilization support for the enhancement of enzyme activity and recyclability is a highly desirable objective. Single-crystalline ordered macro-microporous ZIF-8 (SOM-ZIF-8), has emerged as a highly effective matrix for enzyme immobilization, however, the inherent hydrophobic nature limits its further advancement. Herein, we have customized the immobilization of the Pseudomonas cepacia lipase (LP) in the modification-channels of SOM-ZIF-8 by functionalizing the inner surface-properties with polyethylene glycol (PEG) (LP@SOM-ZIF-8-PEG), and significant enhancement of the activity and (thermal, solvent and cyclic) stability can be realized. The incorporation of PEG into SOM-ZIF-8 regulates its inner surface charge and hydrophobic properties, thereby enhancing enzyme loading, facilitating enzyme conformational adjustments, and achieving a uniform dispersion of LP in SOM-ZIF-8-PEG. LP@SOM-ZIF-8-PEG not only demonstrates a pronounced elevation in enzyme loading and activity over LP@SOM-ZIF-8 but also shows an enzyme activity that is impressively three times greater than LP@ZIF-8. It can completely resolve the 1-phenylethanol racemate in 60 min, with a conversion close to 50 % and an enantioselectivity of 99.8 %. After nine cycles of reuse, the LP@SOM-ZIF-8-PEG still holds onto 95 % of its initial activity. The excellent catalytic performance and stability of LP@SOM-ZIF-8-PEG, along with the universality of the PEG modification strategy for other enzymes, make this work promising in industrial applications.
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Food Bioscience
·
2024
·
IF 4.8
Ji Wang,Xu-Hui Huang,Xusong Wang,Hehe Tian,Liang Wang,Da-Yong Zhou,Lei Qin
Flavor is one of the important sensory attributes of foods. This research explored the effect of lipase on flavor, lipid composition, lipase activity, lipid oxidation, and sensory evaluation of lightly salted large yellow croaker. Results exhibited that the addition of lipase improved the reduced-salt product's flavor. The content of 1-octen-3-ol, nonanal, 2,5-octanedione, hexanal, heptanal, and octanal were key aromas and were increased by the addition of lipase. Through adding the lipase, triacylglycerol (TG) and phosphatidylethanolamine (PE) and could be decompose into fatty acids. Products marinated in 4% NaCl coupled with lipase exhibited higher thiobarbituric acid reactive substance (TBARS) values, acid values, and lipase activity compared with those marinated in 4% NaCl. Moderate promotion of lipid oxidation is beneficial for improving product flavor. Lipase addition had a positive influence on saltiness, aroma, and over-acceptability of reduced-salt products by multi-sensory synergy perception. Especially, addition of 0.2% or 0.4% lipase to reduced-salt products is closest to the flavor profile of normal-salt products. These results provide a new insight into improving reduced-salt foods.
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