HSV在两种细胞培养上抗原出现规律(的观察)及细胞敏感性的比较

本实验观察比较了Hep—2细胞和兔肾细胞对单纯疱疹病毒的敏感性。用免疫荧光法和免疫酶标法检测了细胞培养上HSV抗原出现的规律。实验结果指出:Hep—2细胞和兔肾细胞对HSV的敏感性一致;接种病毒后6小时,可以在Hep—2细胞和兔肾细胞上发现HSV抗原;免疫荧光法和免疫酶标法是进行病毒学研究和病毒感染早期诊断的可靠方法。     

大肠杆菌棉子糖操纵子a-半乳糖苷酶表达的调节控制

大肠杆菌棉子糖操纵子(raf)位于质粒,其第一结构基因rafA编码的a-半乳糖苷酶为诱导酶。Rat操纵子比乳糖操纵子(lac)或蜜二糖操纵子(mel)对诱导物有更严格的结构特异性。该酶被蜜二糖或棉子糖诱导,也被D-半乳糖微弱诱导,但不受乳糖、PNPG等结构相近糖所诱导。A-半乳糖苷酶的酶诱导形成能力在对数生长末期出现高峰。Rat 操纵子基因结构组成及调节与乳糖操纵子相似。以阻遏物为中介的负调控在raf操纵子调节中起主要作用,同时以环腺苷一代谢降解物基因激活蛋白(cAMP-CAP)为中介的正调控也参与调节。当0．4％葡萄糖加入到其它碳源培养基时,该酶表达水平下降至原活力的1／2—1／3。无论诱导或组成型酶的葡萄糖抑制均未见瞬时抑制。腺苷环化酶(cya)缺失或环腺苷受体蛋白(crP)和cya双缺陷菌株的酶表达则分别下降到原活力的9％和2．5％。Cya突变株或葡萄糖对raf操纵子表达的抑制可被cAMP解除,但cya和crP双缺陷菌株仍有葡萄糖抑制,而且这种抑制不为cAMP抵消,表明通过降低cAMp而影响cAMP-CAP复合体形成还不能解释代谢降解物抑制的全部机制。尚无证据说明吲哚类小分子化合物和低浓度尿素对raf操纵子表达的明显作用。     

大肠杆菌棉子糖操纵子α—半乳糖苷酶表达的调节控制

The alpha-galactosidase, coded for by the first structural gene rafA in the plasmid determined raf operon was an inducible enzyme. In contrast to lac or mel operon, raf operon has more strict structural specificity for inducers. The enzyme can be induced by melibiose and raffinose, or weakly by D-galactose, but not by structurally related sugars such as lactose, PNPG etc.. The alpha-galactosidase forming capacity as function of growth curve reached a single peak at the end of the logarithmic phase of the growth. The structure and regulation of raf operon is similar to those of lac operon. The repressormor-mediated negative control plays a major role in the regulation of raf operon, and cAMP-CAP mediated positive control is also involved in the regulation. When 0.4% glucose was added into the medium with other carbon sources, the expression of the enzyme was repressed by 2-3 fold. Transient catabolite repression has been observed neither in inducible nor constitutive alpha-galactosidase expression. Based on alpha-galactosidase assay, in mutant strains CA8306(cya) and CA8445 (cya, crp) the expression level of raf operon was only 9% and 2.5% of that in wild type strain respectively. The glucose effect or the repression in cya mutant can be abolished by 1-5 mmol cAMP. The constitutive alpha-galactosidase expression in cya and cry double mutant (CA8445) remains repressible by glucose, but irreversible by cAMP, suggesting cAMP-CAP complex is not the exclusive mediator of the catablite repression.(ABSTRACT TRUNCATED AT 250 WORDS)     

大肠杆菌青霉素G酰化酶基因及其邻近区域的核苷酸全序列

Some of microorganisms have been known to possess penicillin G acylase activity. The E. coli derived penicillin G acylase (PGA) can catalyze the conversion of penicillin G into phenylacetic acid and 6-amino-penicillanic acid, the latter is used as the starting compound for the industrial formation of semi-synthetic penicillins. Apart from its industrial importance, the enzyme PGA displays a number of interesting properties. Catalytically active enzyme is localized in the periplasmic space of E. coli cells and composed of two dissimilar subunits. The two subunits are apparently produced from a precursor protein, via a processing pathway hitherto unique in its features for a prokaryotic enzyme. The studies on processing of the precursor and on the relationship between structure and function of the mature enzyme are important theoretically. Previously we cloned a 3.5 kb DNA fragment from a strain (E. coli AS 1.76), which displays PGA activity. In this paper, we report a nucleotide sequence of the 3.5 kb DNA fragment containing PGA gene. After insertion of the DNA fragment into EcoR I and Hind III sites in pWR 13, pPGA 20 had been obtained. We subcloned the Hind III and Bg1 II treated fragment of 1.6 kb in length from pPGA 20 into Hind III and BamH I sites of pWR 13 to get a pPGA 1.6, and Bg1 II and EcoR I treated fragment of 1.9 kb in length into BamH I and EcoR I sites of pWR 13 to get a pPGA 1.9. The linearized pPGA 1.9 which were digested with appropriate restriction enzymes were progressively shortened from both ends respectively by digestion with Bal 31 nuclease, followed by cleavage of shortened target DNA off vector DNA molecules with appropriate restriction enzymes. The series of the DNA fragments shortened from EcoR I end were then cloned into plasmid pWR 13 which had previously digested with Hind III and Sma I enzymes (Fig. 1). The DNA fragment cloned in pWR 13 were directly sequenced on the resulted plasmids by using primer I and primer II. Thus we have obtained the complete nucleotide sequence of the 3.5 kb DNA fragment. The 3.5 kb fragment contains an intact PGA gene which is 2.6 kb.(ABSTRACT TRUNCATED AT 400 WORDS)     

酪氨酸对人离体滋养层细胞孕酮与hCG分泌的影响

本文观察三种剂量(2×10~(-5)mol/L,2×10~(-4)mol/L和2×10~(-3)mol/L)的酪氮酸对离体培养的滋养层细胞孕酮及hCG分泌的影响,并对其抑制效应的机理作了初步探讨。实验结果表明,三种剂量的酪氨酸均可抑制滋养层细胞孕酮分泌(P<0.01),但是,在孕酮分泌受酪氨酸抑制的同时,未见对hCG分泌发生影响(P>0.05),进一步观察了酪氨酸对滋养层细胞3β-羟甾脱氢酶活性的影响,结果表明,酪氨酸能显著抑制3β-羟甾脱氢酶活性,提示酪氨酸对滋养层细胞孕酮生成的抑制作用与抑制3β-羟甾脱氢酶活性有关。     

高等植物几丁酶研究进展

几丁醇是一种分解聚N—乙酰胺基葡萄糖分子的糖苷酶,普遍存在于自然界。高等植物中,几丁酶虽然普遍存在,但迄今没有发现底物。因此,高等植物几丁酶的功能是个十分吸引人而又研究不多的问题。本文介绍高等植物几丁酶的分布、酶学性质,基因结构和可诱导性等研究结果。并介绍了几丁酶在植物抗真菌病中的作用的研究进展。     

菠菜叶片过氧体甘氨酸转氨酶初探

菠菜叶片过氧体可转变甘油酸或羟基丙酮酸成为丝氨酸。以甘氨酸为氨基供体时,完整过氧体的转变活性比破碎的约高2.5倍,这不是由于完整的膜包被使有效的羟基丙酮酸浓度增加,或由于膜破碎使辅助因子损失,也不是由于过氧体膜的甘氨酸主动运输系统的作用。结果显示,在过氧体中存在甘氨酸转氨酶,完整过氧体中高的转氨速度可能是由于甘氨酸转氨酶在完整的和破碎的过氧体中的构象或构型不同。     

固定化尖镰孢菌细胞的某些酶学特性

尖镰孢菌(Fusariun oxysporum)33—11是一株青霉素V酰化酶的高产菌株。用二醋酸纤维素固定的该菌细胞裂解青霉素V最适的Ph范围为7．4至7.6;最适的反应温度为45℃至48℃;其酶活性受8-羟基喹啉和EDTA的可逆抑制,Mg²⁺、Zn^2-和Mn^2-离子则有激活作用。采用问歇式搅拌裂解青霉素V,测得0．6rnm颗粒度固定化细胞的表观km值为11．7mmol／L。裂解青霉素V的反应活化能为33kJ／mol;底物抑制常数Ks为1950mmol/L;苯氧乙酸和6-APA的抑制常数分别为220mmol／L和270mm0I／L,在裂解反应中．前者是竞争性抑制剂,后者是非竞争性抑制剂。     

酵母ENO2上游激活顺序对酵母 LEU2表达的影响

本文将酿酒酵母(Saccharomyces cerevisiae)中编码烯醇化酶的基因之—ENO2的上游墩活顺序嵌入穿梭质粒YEpl3上酵母LEU2基因上游—405Hpa I的酶切部位。LEU2为编码β-异丙基苹果酸脱氢酶基因。从而研究了酵母ENO2的上游激活顺序对酵母LEU2表达的影响。实验结果表明ENO2上游激活顺序不论正向或反向嵌八都激活LEU2的表达达四倍左右。有亮氨酸存在的条件下,LEU2的表达受到抑制。ENO2上游激活顺序在对LEU2表达的激活上并不受葡萄糖的诱导。提出了用ENO2上游激活顺序组建高表达系统的可能性。