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1.
化学修饰木瓜蛋白酶的固定化及性质研究 总被引:1,自引:0,他引:1
在底物保护和无底物保护下,用丁二酸酐对木瓜蛋白酶进行化学修饰,以三硝基苯磺酸法测定修饰酶的平均氨基修饰度,以棉布为载体,戊二醛为交联剂,对修饰前后的木瓜蛋白酶分别进行固定化.考察了温度、pH和表面活性剂SDS对化学修饰的固定化木瓜蛋白酶活力的影响,并与固定化天然木瓜蛋白酶进行了比较.研究表明,化学修饰固定化木瓜蛋白酶的最适反应温度为80℃;最适pH为9.0;在SDS浓度为20mg/mL时酶活也仍能保持在40%左右;米氏常数为187g/L.与天然的固定化酶相比,化学修饰的固定化木瓜蛋白酶的热稳定性、耐碱性和耐洗涤性得到了显著提高. 相似文献
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氨化大孔球状聚氯乙烯固定化木瓜蛋白酶的研究 总被引:9,自引:0,他引:9
以氨化大孔球状聚氯乙烯为载体,采用戊二醛载体交联的方法,将木瓜蛋白酶进行了固定化。以酪蛋白为底物,测定了固定化酶的活力回收。研究了固定化条件对固定化酶活力回收的影响。同时,对所得固定化酶的性质,如温度-活力关系、pH-活力关系、热稳定性以及重复使用性进行了考察。结果表明,所得固定化木瓜蛋白酶具有较好的稳定性和重复使用性。 相似文献
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大孔阴离子树脂DEAE-E/H固定化氨基酰化酶的研究 总被引:7,自引:1,他引:6
以弱碱性大孔阴离子树脂DEAE-E/H为载体固定化氨基酰化酶.通过对影响固定化结果的几个因素,如树脂的离子类型、pH值、温度,以及自由酶液浓度等进行系统研究,得到了适宜的固定化条件:将DEAE-E/H转化为Ac-型;自由酶液浓度120U/ml,pH值6.5;固定化温度为常温.在此条件下制备的固定化氨基酰化酶比酶活可达1200U/g~1500U/g,酶活保留率超过60%.DEAE-E/H作为固定化载体,具有价格低廉,物理性能好,固定化方法简便等特点,具有很好的工业应用前景. 相似文献
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聚丙烯酰胺固定化糖化酶特性的研究 总被引:5,自引:0,他引:5
本研究以丙烯酰胺单体通过反向悬浮聚合技术合成聚丙烯酰胺作为载体材料,采用包埋—交联法固定化葡萄糖淀粉酶,并对其特性进行了研究.结果表明,该固定化酶最适pH值为5.0,最适温度为55~58℃,而且具有较好的贮存稳定性和操作稳定性,8个月后该固定化酶的残余活力仍保持在94%左右,可重复使用43批次,此固定化酶酶活回收率达到56%.实验表明丙烯酰胺悬浮聚合固定化糖化酶的方法是简便可行的. 相似文献
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漆酶在纳米多孔金上的固定化及其酶学性质研究 总被引:1,自引:0,他引:1
利用纳米材料为载体对酶等生物大分子进行固定化近年来引起人们的浓厚兴趣. 以Au/Ag合金为原料, 通过控制浓硝酸的腐蚀时间再辅以退火处理得到了不同孔径的纳米多孔金(NPG), 利用扫描电镜(SEM)和N2气体吸附仪对孔性质进行了表征. 以NPG为载体, 用α-硫辛酸和N-乙基-N’-(3-二甲基氨基丙基)碳酰二亚胺/N-羟基琥珀酰亚胺(EDC/NHS)对金表面进行活化, 通过化学共价偶联的方法对产自Trametes versicolor的漆酶进行了固定化. 比较了孔径大小对酶固定化量及比活力的影响. 发现小孔径更有利于对该漆酶的固定化. 与游离酶相比, 固定化酶的最适pH没有改变, 但最适温度却从原来的40 ℃升到了60 ℃. 固定化后, 漆酶的pH和热稳定性都明显提高了. 重复使用8次仍能保持初始活力的65%, 且在4 ℃下保存1个月几乎观察不到酶活力的下降. 此外, 失活的固定化酶经浓硝酸处理后, NPG载体可重复利用. 本结果初步显示出了NPG在生物技术领域中的应用潜力. 相似文献
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甲醛活化壳聚糖制备固定化l—天冬酰胺酶 总被引:12,自引:0,他引:12
研究了以甲醛活化的壳聚糖为载体固定化l-天冬酰胺酶,其活力回收可达到113%,比用以戊二醛为交联剂进行固定化的活力回收要高出五倍多。分别从缓冲液的pH值及甲醛用量等方面进行了固定化l-天冬酰胺酶的条件优化,并且对固定化酶的性质作了探讨。 相似文献
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HPD-750树脂是中极性大孔吸附树脂,生物相容性好,机械性能稳定,具有较大的比表面积,可用于固定化酶载体材料。本文以HPD-750大孔树脂为载体固定化果胶酶,研究各因素对固定化酶的影响,并采用正交试验对固定化条件进行优化。结果表明,当pH为4.0、固定化温度为45℃、固定化时间为4h、加酶量为0.16g/mL时,固定化酶活力可达5146U/mg。以HPD-750大孔树脂为载体材料制备的固定化酶相较于游离酶具有更好的酸碱稳定性和热稳定性。在循环使用10次后,酶活力依然保留80%以上;4℃储藏25d之后,其酶活力仍保留60%以上。与D311大孔树脂、聚丙烯酰胺和海藻酸钠微球制备的固定化酶相比,HPD-750大孔树脂固定化酶的活性、操作稳定性、机械稳定性和储存稳定性都较好。该结果说明,HPD-750大孔树脂可作为固定化酶较好的载体材料。 相似文献
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酸性接枝淀粉固定化糖化酶及稳定性研究 总被引:3,自引:0,他引:3
糖化酶(EC.3.2.1.3,GA)是工业上应用规模最大的3种酶制剂之一.目前,工业生产中大多采用游离GA,这给产物的分离和纯化带来诸多不便.因此,采用固定化糖化酶(IGA)是酶制剂工业发展的必然趋势. 相似文献
12.
Mohy Eldin MS Seuror EI Nasr MA El-Aassar MR Tieama HA 《Applied biochemistry and biotechnology》2011,164(1):10-22
ρ-Benzoquinone-activated alginate beads were presented as a new carrier for affinity covalent immobilization of glucoamylase
enzyme. Evidences of alginate modification were extracted from FT-IR and thermal gravimetric analysis and supported by morphological
changes recognized through SEM examination. Factors affecting the modification process such as ρ-benzoquinone (PBQ) concentration,
reaction time, reaction temperature, reaction pH and finally alginate concentration, have been studied. Its influence on the
amount of coupled PBQ was consequently correlated to the changes of the catalytic activity and the retained activity of immobilized
enzyme, the main parameters judging the success of the immobilization process. The immobilized glucoamylase was found kept
almost 80% of its native activity giving proof of non-significant substrate, starch, diffusion limitation. The proposed affinity
covalent immobilizing technique would rank among the potential strategies for efficient immobilization of glucoamylase enzyme. 相似文献
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M. G. Roig A. Slade J. F. Kennedy D. W. Taylor M. G. Garaita 《Applied biochemistry and biotechnology》1995,50(1):11-33
The covalent immobilization of glucoamylase on new epoxide-, isocyanate-, acid chloride-, and carboxylic acid-activated plastic
supports shows the viability of such supports for immobilizing enzymes (especially those reacting with 1,6-diaminohexane and
glutaraldehyde) for producing side arms. The operational stability of immobilized glucoamylase could be extended by crosslinking
the enzyme, by increasing the substrate concentration, or by extending the support’s side arm. The pH curves for the immobilized
enzyme were in general not found to be shifted from the pH optimum of the soluble enzyme. However, the immobilized enzyme’s
temperature activity profiles were shifted to a lower temperature range when compared to the soluble enzyme. The immobilized
glucoamylase Michaelis constants increased, and the maximum rates and specific activities decreased with respect to the soluble
enzyme kinetic parameters. 相似文献
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Glucoamylase was immobilized onto novel porous polymer supports. The properties of immobilized glucoamylase and the relationship
between the activity of immobilized enzyme and the properties of porous polymer supports were investigated. Compared with
the native enzyme, the temperature profile of immobilized glucoamylase was widened, and the optimum pH was also changed. The
optimum substrate concentration of immobilized glucoamylase was higher than that of native enzyme. After storage for 23 d,
the immobilized glucoamylase still maintained about 84% of its initial activity, whereas the native enzyme only maintained
about 58% of the initial activity. Moreover, after using repeatedly seven times, the immobilized enzyme maintained about 85%
of its initial activity. Furthermore, the properties of porous polymer supports had an effect on the activity of the immobilized
glucoamylase. 相似文献
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We present a novel type of nanoreactor suitable for the immobilization of enzymes. The particles used consist of a polystyrene core onto which long chains of poly(acrylic acid) are grafted ("spherical polyelectrolyte brush"). Proteins adsorbed spontaneously onto these particles from aqueous solutions if the ionic strength is low. We immobilized glucoamylase on these particles and showed that this enzyme keeps nearly its full activity. This is shown by analyzing the enzymatic activity in terms of the Michaelis-Menten kinetics. No leaching out of the enzyme takes place during the reaction and the colloidal stability is not impeded by the adsorbed biomolecules. The data presented here suggest that the principle of immobilizing enzymes on these particles may be of general use.The Figure shows a schematic representation of the colloidal nanoreactors. 相似文献
17.
A novel affinity covalent immobilization technique of glucoamylase enzyme onto ρ-benzoquinone-activated alginate beads was
presented and compared with traditional entrapment one. Factors affecting the immobilization process such as enzyme concentration,
alginate concentration, calcium chloride concentration, cross-linking time, and temperature were studied. No shift in the
optimum temperature and pH of immobilized enzymes was observed. In addition, K
m values of free and entrapped glucoamylase were found to be almost identical, while the covalently immobilized enzyme shows
the lowest affinity for substrate. In accordance, V
m value of covalently immobilized enzyme was found lowest among free and immobilized counter parts. On the other hand, the
retained activity of covalently immobilized glucoamylase has been improved and was found higher than that of entrapped one.
Finally, the industrial applicability of covalently immobilized glucoamylase has been investigated through monitoring both
shelf and operational stability characters. The covalently immobilized enzyme kept its activity over 36 days of shelf storage
and after 30 repeated use runs. Drying the catalytic beads greatly reduced its activity in the beginning but recovered its
lost part during use. In general, the newly developed affinity covalent immobilization technique of glucoamylase onto ρ-benzoquinone-activated
alginate carrier is simple yet effective and could be used for the immobilization of some other enzymes especially amylases. 相似文献
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Summary The enzymes a-amylase, invertase and glucoamylase were immobilized on acid activated montmorillonite using two techniques,
viz. adsorption and covalent binding, and their activities were tested in a batch and packed-bed reactor and were compared. The
packed-bed reactor showed an improved performance for all immobilized enzymes, which was attributed to lowering of diffusional
restrictions to mass transfer. Lower activity in case of batch reactor for immobilized invertase was due to a combined effect
of loss of native conformation of enzyme on account of immobilization and mass transfer resistances due to improper diffusion
of substrate to the active site of enzyme. For immobilized glucoamylase, the packed-bed reactor demonstrated exceptionally
high activity that was very close to the free enzyme. Covalently bound glucoamylase showed higher activity than the free enzyme. 相似文献
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Jia Xian HUANG* Yan Li HUO Yue LI Zhi YUAN Department of Chemistry the State Key Laboratory of Functional Polymer Materials for Adsorption Separation Nankai University Tianjin 《中国化学快报》2001,(11)
Usually, before enzyme was immobilized onto support materials, these support materials had been activated through some activators, such as glutaraldehyde. The glucoamylase has been covalently immobilized onto several different support materials through the formation of Schiff base1-5. In this work, the glucoamylase was covalently (in the form of σ-bond) immobilized onto the porous polymer supports containing cyclic carbonate without activation. The relationship between the activity of the … 相似文献
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Steven J. Swanson Alden Emery Henry C. Lim 《Applied biochemistry and biotechnology》1976,1(2):119-135
The activity of immobilized enzymes decays with time, and the capacity of a carrier will decrease with repeated regeneration. Relations between production cost and these factors are shown, and demonstrated with data on glucoamylase immobilized on porous glass. Optimum design calls for very low temperature and for cycle times several years long. A practical design may be made by limiting cycle time to an upper limit and calculating the temperature for which this time is optimum. In this case, reagents and carrier are the most important costs, even with an expensive enzyme. 相似文献