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以壳聚糖为载体固定化青霉素酰化酶的研究 总被引:12,自引:1,他引:11
介绍了以壳聚糖为载体固定化青霉素酰化酶需首先制备壳聚糖颗粒 ,使用戊二醛、甲醛、乙二醛 3种活化剂处理所得的壳聚糖颗粒 ,确定了以戊二醛为活化剂交联其上的氨基共价结合青霉素酰化酶的固定化方法。从戊二醛的浓度、pH值、固定化时间、固定化pH值、酶用量等条件摸索了最佳固定化条件 ,获得了酶活力为4 0 0 0 0U/ (g·h)、回收率为 5 0 %左右的固定化青霉素酰化酶。 相似文献
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青霉素酰化酶又称为青霉素酰胺酶或青霉素氨基水解酶,主要从大肠埃希菌胞内酶和巨大芽孢杆菌胞外酶获得,该酶已大规模应用于工业生产β-内酰胺类抗生素的关键中间体和半合成β-内酰胺类抗生素。主要介绍了青霉素酰化酶固定化技术的进展,讨论了不同固定化技术的特点,并展望了固定化青霉素酰化酶的发展前景。 相似文献
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在反应温度为45℃、反应时间为6 h、丙烯酰胺(AM)质量浓度为40 g/L、引发剂用量为单体质量1%的条件下制得PAM-SiO2复合载体,利用红外光谱表征了其化学结构,热失重法测定其接枝量为21%。在所制PAM-SiO2复合载体上固载青霉素酰化酶,其固载最适条件为:戊二醛用量0.1 mmol/L,pH值7.64,给酶量1%,温度42℃,时间11 h~12 h。此条件下所得固定化酶的活力为34000 U/mL;测得所制固定化酶的最适pH值为7.82,最适温度为45℃。 相似文献
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利用共沉淀方法制备出饱和磁化强度为66.2emu/g的Fe3O4,通过溶胶-凝胶进行氨基功能化改性后饱和磁强度仍高达27.2e-mu/g,其剩磁和矫顽力均为0。对不同反应工艺制备的磁性复合载体进行固定化青霉素酰化酶(PGA),当戊二醛的浓度为0.02%时,固定化酶的起始表观酶活为3198.38 IU/g,当戊二醛的浓度为0.1%时,进行持续间歇8次使用后,活性为起始活性的86.1%。 相似文献
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磁性微粒对青霉素G酰化酶的固定化研究 总被引:1,自引:0,他引:1
将无机磁性粒子Fe3O4与有机材料海藻酸钠结合起来制成一种复合的磁性微粒,并将其进行表面修饰,通过化学共价法来固定青霉素G酰化酶。通过扫描电镜等对微粒进行形态学观察,并用傅立叶红外图谱表征微粒表面修饰基团。酶学性质研究表明,该微粒固定化的青霉素G酰化酶的最适pH值为7.5,最适温度为40℃。固定化酶与底物的亲和力有所降低,但是稳定性显著提高。重复催化研究结果表明,固定化酶具有比游离酶更广泛的温度及pH值适用范围,并且具有良好的热稳定性、可循环使用性和贮存稳定性。 相似文献
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介绍了青霉素酰化酶的生产及应用新进展。着重介绍了青霉素酰化酶固定化技术的发展。青霉素酰化酶主要应用于6氨基青霉烷酸的工业生产和半合成的β内酰胺抗生素的合成,是在半合成抗生素的生产上有重要作用的一种酶。此外,青霉素酰化酶也可应用于其它的生物转化,如肽的合成、手性化合物的外消旋混合物的拆分。 相似文献
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Functionalized ionic liquids containing ethyoxyl groups were synthesized and immobilized on magnetic silica nanoparticles(MSNP) prepared by two steps,i.e.,Fe3O4 synthesis and silica shell growth on the surface.This magnetic nanoparticle supported ionic liquid(MNP-IL) were applied in the immobilization of penicillin G acylase(PGA).The MSNPs and MNP-ILs were characterized by the means of Fourier transform infrared spectroscopy(FTIR),scanning electron microscopy(SEM),transmission electron microscopy(TEM),and vibrating sample magnetometer(VSM).The results showed that the average size of magnetic Fe3O4 nanoparticles and MSNPs were ~10 and ~90 nm,respectively.The saturation magnetizations of magnetic Fe3O4 nanoparticles and MNP-ILs were 63.7 and 26.9 A?m2?kg?1,respectively.The MNP-IL was successfully applied in the immobilization of PGA.The maximum amount of loaded enzyme was about 209 mg?g?1(based on carrier),and the highest enzyme activity of immobilized PGA(based on ImPGA) was 261 U?g?1.Both the amount of loaded enzyme and the activity of ImPGA are at the same level of or higher than that in previous reports.After 10 consecutive operations,ImPGA still main-tained 62% of its initial activity,indicating the good recovery property of ImPGA activity.The ionic liquid modified magnetic particles integrate the magnetic properties of Fe3O4 and the structure-tunable properties of ionic liquids,and have extensive potential uses in protein immobilization and magnetic bioseparation.This work may open up a novel strategy to immobilize proteins by ionic liquids. 相似文献
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To improve the covalent immobilization of penicillin acylase (PA), macromolecular crowding theory was applied to its immobilization. Influence of mass ratio of enzyme to the silica, as well as, activation time with glutaraldehyde on the activity of assembled PA, was studied. In the mesopores, the effect of fl-cyclodextrin (β-CD) on the immobilization of the enzyme was also investigated. It was remarkable that the coupled yield and relative activity reached 99.5% and 92.3%, respectively, when penicillin acylase assembled covalently in the mesopores. The results here indicate that mimicked macromolecule crowding could significantly ameliorate the performance of covalently immobilized PA. 相似文献
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通过溶剂热法制备出高磁性的聚合物微球MnFe_2O_4,经正硅酸乙酯(TEOS)和3-氨基丙基三乙氧基硅烷(APTS)对微球表面进行改性修饰,制备出新型复合材料MnFe_2O_4@SiO_2-NH_2,并将其用于固定青霉素G酰化酶。在Si/Fe比为7 mmol/g、n(TEOS)∶n(APTS)=1∶1时,固定化酶PGA/MnFe_2O_4@SiO_2-NH_2在37℃下水解青霉素G钾合成6-氨基青霉烷酸,表观酶活为1 660 IU/g、载酶量为107.1 mg/g、比酶活为15.5 IU/mg、活性回收率为46.9%。经过6次重复使用,保留初始酶活的81.3%,在使用中固定化酶在磁场的作用下能够快速沉降与产物分离。 相似文献
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反胶束体系中青霉素酰化酶活性及稳定性研究 总被引:6,自引:0,他引:6
反胶束体系是一种新型的酶反应介质体系。构建合适的反应体系,使青霉素酰化在该体系中保持较高的酶活性,是实现反胶束体系中酶法合成头孢类抗生素的关键所在。本文分别考察了青霉素酰化酶在CTAB/正己醇和CTAB/异辛烷/正己醇两种反应胶束体系中不同水含量和pH值对酶活力的影响。结果表明:包埋于CTAB/异辛烷/正己醇反胶束体系中的青霉素酰化酶在W0为14,pH为7.2时表现出较高的酶活性。试验中还考察了不同温度对包埋于CTAB/异辛烷/正己醇反胶束体系中的青霉素酰化酶活力的影响,并与游离酶液进行了比较,该酶在所采用的反胶束体系中的最适水解反应温度为39℃,较游离酶液的最适水解温度升高了2℃。同时酶的热稳定性亦较游离酶液有所提高,表现在CTAB/异辛烷/正己醇反胶束体系中,经50℃保温240min,酶活保留率为76.0%,而在同样条件下游离酶液仅为36.5%。 相似文献
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报道了青霉素酰化酶法制备D-丙氨酸的研究结果,为D-丙氨酸的制备提供了一种方法。考察了pH、温度、DL-丙氨酸与苯乙酰氯的摩尔比、反应时间对制备结果的影响。确定了制备N-苯乙酰-DL-丙氨酸的最佳反应条件:pH=9.5,n(DL-丙氨酸)∶n(苯乙酰氯)=1.1∶1;酶促反应最佳条件为:pH=7.5,温度37℃条件下反应6h,其产物为N-苯乙酰-D-丙氨酸和苯乙酸的混合物。该混合物用c(HC l)=6 mol/L的盐酸高温回流6 h得D-丙氨酸盐酸盐,再经717阴离子树脂处理得D-丙氨酸。光学纯度为95.3%,总收率为72.5%。 相似文献
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Dr. Chengcheng Tian Xuyang Xu Haiyang Zhang Dr. Nada Mehio Dr. Yanglong Guo Dr. Liang Ma Dr. Sheng Dai 《Chembiochem : a European journal of chemical biology》2020,21(15):2143-2148
Penicillin G acylase (PGA) has been immobilized on a lanthanum-incorporated mesostructured cellular foam (La-MCF) support by using the interaction between the strong Lewis acid sites on the surface of La-MCF and the free amino groups of lysine residues of PGA. The La-MCF support was successfully synthesized in situ through the addition of a citric acid (CA) complexant. The results of pyridine-IR spectroscopy show the presence of strong Lewis acid sites on the surface of the prepared La-MCF (with CA), attributed to the incorporation of lanthanum species into the framework of MCF. Through interaction with the strong Lewis acid sites, the enzymes can be firmly immobilized on the surface of the support. The results indicate that PGA/La-MCF (with CA) exhibits a high specific activity and greatly enhanced operational stability. For the hydrolysis of penicillin G potassium salt, the initial specific activity of PGA/La-MCF (with CA) reaches 10023 U/g. Even after being recycled 10 times, PGA/La-MCF (with CA) retains 89 % of its initial specific activity, much higher than the 77 % of PGA/Si-MCF. 相似文献
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对产青霉素G酰化酶的温度诱导型重组枯草芽孢杆菌发酵条件进行了研究。结果表明,培养基的最佳氮源和碳源组成为:35gL-1牛肉膏、3gL-1葡萄糖和6gL-1淀粉;最佳诱导时机是细胞对数生长的中后期,诱导前应将pH调节至中性;最佳诱导条件为升高温度至50℃并维持4min;随后将温度降低到34℃进行重组蛋白的表达。在上述条件下,PGA的表达水平可以达到5.8UmL-1。SDS-PAGE电泳分析表明该重组菌能够将绝大部分表达的PGA分泌到细胞外,而且表达的PGA蛋白占有高比例(90%)。 相似文献