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1.
根据文献报道的核苷酸序列合成Bacillus deramificans普鲁兰酶成熟肽编码基因BdP。将BdP基因插入芽孢杆菌分泌表达载体pUC980信号肽编码区下游,获得重组质粒pUC980-BdP,重组质粒转化中温α-淀粉酶生产菌解淀粉芽孢杆菌BF7658菌株。摇瓶发酵实验表明,重组转化子发酵液有明显普鲁兰酶酶活,约48h酶活达到最高水平,为2.8ASPU/mL。酶学性质分析表明,重组酶最适作用温度约为60℃,最适反应pH为5.0,60℃保温3h仍保存50%的活性。重组酶性质适合淀粉糖化工艺的要求。  相似文献   

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目的:对枯草芽孢杆菌TM903嘌呤核苷磷酸化酶进行分离纯化及酶学性质研究。方法:经加热、硫酸铵盐析和SephadexG-100凝胶过滤,对枯草芽孢杆菌TM903中的嘌呤核苷磷酸化酶进行分离纯化,并对其酶学性质进行研究。结果:酶的最适反应温度为65℃,最适反应pH值为7.5,在30-50℃时热稳定性较好;K^+对该酶有激活作用,而Na^+、ca^+、Mg^+、Mn^+等金属离子对该酶有抑制作用;Km值为2.11mmol/L,Vmax值为0.84mmol/(min·L)。结论:分离纯化了枯草芽孢杆菌TM903嘌呤核苷磷酸化酶,并研究了其酶学性质,为利巴韦林的发酵工艺优化提供了重要的酶学理论基础。  相似文献   

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目的:对Anoxybacillussp.DL3的产蛋白酶条件及其酶学性质进行研究,为下一步进行蛋白酶基因的克隆、表达提供依据。方法:应用常规方法液体培养细菌,研究温度、pH、培养基中碳源、氮源对菌株产蛋白酶的影响,硫酸铵盐析的方法提取酶液,并采用Folin法测酶活性。用紫外分光光度计在OD680hi/1测吸光值。并对提取的蛋白酶液进行酶的最适温度、pH以及酶的热稳定性和pH稳定性研究,向酶液中添加金属离子和EDTA、PMSF,研究其对酶活性的影响。结果:在培养基初始pH是6.5,培养温度为40℃时菌株产酶酶活性最大;培养基中以乳糖为碳源,酵母膏和硫酸铵为氮源,碳源与氮源的比例为1:2时,酶活最大。酶学性质研究结果显示:该酶的最适反应温度是55℃,最适反应pH是7.0;在50℃保温20min-80min内,酶活力下降幅度较小。60℃保温60min后,仍保持约60%的酶活。70℃保温60min后,残余酶活为30%。该酶在pH为6.0~8.0范围内,相对酶活差别不是很大,下降趋势大致相同。在强碱条件下,相对酶活下降很明显。Fe2+、Cu2+和Hg2+对酶活性有明显的抑制作用;Ca2+、Mg^2+、Mn2+等金属离子对酶活性有明显的促进作用;乙二胺四乙酸(EDTA)和苯甲基磺酰氟(PMSF)对酶活性也有一定抑制作用。结论:Anoxybacillussp.DL3所产的蛋白酶为嗜热中性蛋白酶,此酶具有较好的热稳定性和pH耐受性,该菌株具有进一步开发、利用的价值。  相似文献   

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木聚糖酶产生菌的选育及发酵条件研究   总被引:17,自引:2,他引:15  
采用富集培养、透明圈平板筛选,从长期在其上堆放秸秆的土壤中分离到-株木聚糖酶产生菌黑曲霉C2。对该菌株进行紫外线诱变处理,然后采用自然选育的方法筛选到-株木聚糖酶高产菌株C2—56,并对该菌株的适宜发酵条件和酶学特性进行了初步研究。结果表明,C2—56三角瓶发酵酶活达656.8U/g,比出发菌株C2提高了140%;发酵条件以80%麸皮和20%玉米芯为培养基,采用3%接种量,在曲盘上发酵72h为宜,酶活达1750U/g;该菌株产生的木聚糖酶具有较差的热稳定性和较好的pH稳定性,适宜反应温度和pH分别为50℃和pH4.2。  相似文献   

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Aspergillus sp.脂肪酶发酵条件优化及酶学性质的研究   总被引:2,自引:0,他引:2  
作者为了得到一种热稳定性较好的脂肪酶新酶种,通过研究分离白极端环境的Aspergillus sp.的最佳产酶条件及其所产脂肪酶的酶学性质,得出了该菌产酶的最佳发酵条件为:以1%黄豆饼粉为氮源、0.2%玉米淀粉为碳源,1.5%橄榄油为诱导物,起始pH6.0左右。装量10mL(250mL三角瓶。摇瓶转速180r/min)、发酵时间为96h。在最佳发酵条件下可得最大发酵酶活36U/mL。Aspergillus sp.所产的脂肪酶的酶学性质是:最适pH为9.0,在pH5.0—10.0于20℃下放置24h后,残余酶活仍保持在起始酶活的90%以上;该酶的最适温度为50℃,50℃保温60min后仍保留70%以上的酶活。Aspergillus sp.所产脂肪酶的热稳定性较好。  相似文献   

6.
壳聚糖固定化德氏根霉脂肪酶的研究   总被引:4,自引:0,他引:4  
研究了壳聚糖吸附和戊二醛交联对脂肪酶固定化条件,在室温条件下将0.4g酶粉溶于pH6.0缓冲液中,加入10g壳聚糖,摇匀,再加入浓度为0.6%戊二醛交联6h,得到固定化酶,酶活力回收率约为54.2%。固定化酶的半失活温度比游离酶的高,半失活温度由游离酶的47℃提高到100℃,最适反应温度由40℃上升至80℃,最适pH由6下降到5.5,固定化酶K’m值由游离酶的Km 50mg/mL增加到56mg/mL。该固定化脂肪酶用于酯的合成;在80℃条件下经过10批次连续水解植物油反应,固定化酶的活力仍保持在82.6%以上。  相似文献   

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嗜热芽孢杆菌XJT—9503菌的发酵液经硫酸铵和丙酮分级沉淀分离纯化得到聚丙烯酰胺凝胶电泳均一的高温中性蛋白酶制品。SDS—PAGE测得酶分子量为3000。当以酪蛋白为底物时,酶反应最适温度为65℃,最适pH为7.在PH6.5—9范围内稳定。在65℃、0、02MpH7.5的磷酸缓冲液中的半衰期为54min。金属离子铜、汞、铝强烈抑制酶活,钙离子,镁离子对酶活有促进作用。  相似文献   

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对重组荧光素酶大肠杆菌菌株M15/pQE30-luc进行了表达条件的优化研究。单因素结果表明:在初始pH值7.0,装液量为20%,2%的接种量,终浓度为0.5mmol/L的IPTG,添加10—30mmol/L的Mg^2+,摇床转速为200r/min,37℃诱导3.5h酶的表达量最高。正交试验结果表明:初始pH值为7.0,添加40mmol/LMg^2=,接种量2%,装液量为20%时表达量最高,比酶活达1.63×10^8RFU/mg蛋白。  相似文献   

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芽孢杆菌EZ菌在55C下生长良好.在培养液中能大量积累胞外纤维素酶(190m㈣/L培养液),所产生的纤维素酶为单一的羧甲基纤维素酶((CMCase)。羧甲基纤维素钠(CMC—Na)为其专一性底物。该酶作用的pH为4.5 8.0;最适pH为6.5 7.0;在pH4.0- 8.0范围内较稳定。酶作用的最适温度为55C、;在60C、处理 10、30、60、90以及120分钟后,残余酶活分别为95%、80.3%、41.4%,19.3%和7.0%;在65C和70C处理10分钟后残余酶活分别为59%N 19%。M2+、Ca2+对酶促反应稍有促进作用;Ag-、Mn2-、Cu2-、Fe2-和Zn2+有明显的抑制作用。  相似文献   

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从腐烂的菊芋及实验室保存的菌种中,选育到一株发酵菊芋产乙醇的菌株克鲁维酵母Kluyveromyces marxianus Y1。利用正交实验法对克鲁维酵母产菊粉酶的培养基组成及培养条件进行优化,确定培养基组成(g/L)为:菊粉40,酵母粉4,蛋白胨4,尿素1;初始pH5.0,温度30℃,150r/min条件下培养达到最佳产酶效果(57U/mL)。该菌株所产菊粉酶的性质测定结果表明:以菊粉为底物,该菊粉酶最适反应温度为55℃,在60℃以下稳定性很好,高于60℃时酶迅速失活;最适pH为5.0,pH4.6—5.2范围内酶稳定性很好;该酶属于外切型菊粉酶,体积分数为8%的乙醇对酶活力基本没有影响。  相似文献   

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K. Hausmann 《Protoplasma》1979,100(2):199-213
Summary The membranes of the pellicle of the ciliatePseudomicrothorax dubius are investigated using thin section electron microscopy and freeze-fracture replicas. The plasma membrane is covered by a surface coat and is connected to the outer alveolar membrane by short, sometimes branched, bridges. The inner alveolar membrane is coated on both sides. The epiplasm lies in intimate contact with the cytoplasmic surface of this membrane, and there is a corresponding deposit on the other surface. This deposit is regularly striated.The epiplasmic layer and the alveoli are interrupted at sites of cytotic activity,e.g., the attachment sites of trichocysts, the cytoproct, and the parasomal sacs. The striated deposit ends where the epiplasm ends, indicating a direct relationship between these two epimembranous layers.There is a deposit along the sides of the first part of the tip of the trichocysts, and in this region the trichocyst membrane is free of intramembranous particles.The membrane of the parasomal sacs has a coat on both surfaces. That on the extraplasmic surface is similar to the surface coat of the plasma membrane. The origin of the cytoplasmic coat is unknown. The cytotic activity of these sacs is indicated by their highly irregular profiles.  相似文献   

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Summary The differentiation of the spermatid, especially in reference to the formation of the flagellum, and transformation of the shape of the nucleus was investigated in the domestic fowl.In the early stage of the spermatid, a prominent Golgi apparatus appears around the centrioles. The Golgi vesicles then surround the axial-filament complex which develops from the distal centriole. These vesicles fuse to form continuous membrane at the earliest stage of flagellar formation, and in the succeeding stage Golgi lamellae are attached to the plasma membrane of the developing flagellum. From these observations, it is assumed that Golgi apparatus may be a source of the membrane system of the flagellum.The microtubules distributed around the nucleus form the circular manchette. The anterior region of the nucleus with the manchette is cylindrical in shape and the posterior region without it remains irregular in shape. When the circular manchette has been completed, the whole nucleus acquires a slender cylindrical shape. The circular manchette then changes into the longitudinal manchette. The nuclei of spermatids without a longitudinal manchette are abnormal in shape. In view of these observations it is assumed that the nuclear shaping of the spermatid may be accomplished by circular manchette and the maintenance of shape of the elongated nucleus by longitudinal manchette.The authors wish to thank Mr. Takayuki Mori for his helpful suggestions and technical advices  相似文献   

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Summary The choriocapillaris is a fenestrated capillary bed located posterior to the retinal pigment epithelium. It serves as the main source of supply to the photoreceptors, retinal pigment epithelium, and other cells of the outer retina. The permeability of these capillaries to intravenously injected ferritin (MW — approx. 480,000; mol. diam. 11 nm) was examined in the mouse, rabbit, and guinea pig, each of which is characterized by a different type of retinal vascularization. In all three species, the bulk of the ferritin remained in the capillary lumina, where it appeared to be blocked at the level of the diaphragmed fenestrae. Some ferritin was present in endothelial cell vacuoles. The results confirm previous work on the rat choriocapillaris and indicate that the barrier function of the choriocapillary endothelium is present even among species in which the retinal circulation differs significantly.Supported by NIH grant EY03418  相似文献   

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This study aimed to analyze the aqueous humor (AH) and the vitreous body (VB) of the eye of the adult frog Rana temporaria L. as a representative species of amphibians, which lead a semi-terrestrial life. The presence of collagen, albumin, uric acid and electron donors was shown in both media; however, there are slight differences in their concentrations. To determine collagen, a spectral-fluorescent probe, cyanine dye, was used. The presence of collagen in AH of the frog was found at the first time. The total content of electron donors (ascorbic and uric acids, tryptophan, and tyrosine) in VB and HA was roughly estimated at ~ 1.5 × 10− 4 mol/L. Both VB and AH absorb light in similar UV regions. The total protein and albumin contents in AH were found to be somewhat higher than those in VB. The uric acid content was at an equally low level in both intraocular media. It is supposed that the similarity of VB and AH compositions shown in this work is due to some exchange between VB and AH contents in the course of accommodation. The role of intraocular fluids in physiological functions of the eye and in protecting the retina against UV light is discussed.  相似文献   

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