L-缬氨酸对环孢菌素形成的影响

环孢菌素的生物合成路线受添加外源氨基酸影响。Ｋｏｂｅｌ和Ｔｒａｂｅｒ^［１］报道过,添加与环孢菌素形成有关的特殊氨基酸可以提高环孢菌素某组分所占的比例。如ＤＬ-氨基丁酸能够提高环孢菌素A的产量,而抑制其它环孢菌素组分产生。本文报道添加L-和DL-缬氨酸对环孢菌素的主要成分环孢菌素A形成的影响。     

雪白白僵菌红色素的初步鉴定及其与环孢菌素A对底物缬氨酸的竞争

【目的】分离鉴定雪白白僵菌红色素,分析红色素与环孢菌素A对底物缬氨酸的竞争和相互影响。【方法】对红色素进行分离纯化,利用UV、IR和ESI-MS对红色素进行初步鉴定。采用缬氨酸分批补料培养,通过控制溶氧水平,以及添加红色素,分析红色素与环孢菌素A合成之间的竞争关联及相互影响。【结果】经鉴定,雪白白僵菌红色素分子式为C15H10O5,推测为含有芳环结构的蒽醌类化合物。在补加缬氨酸和高DO条件下,环孢菌素的产量高于低DO水平,相反红色素在低DO条件下合成量大于高DO水平。在不补加缬氨酸条件下,实验结果与补加缬氨酸培养一致,但是红色素和环孢菌素A的产量都显著降低。进一步添加外源纯化的红色素时,随着添加量的增加出现了环孢菌素A合成先减弱后增加的变化。【结论】发现并证实了雪白白僵菌红色素与环孢菌素A合成都以缬氨酸为共同底物,但两者的途径又相互独立。     

大孔吸附树脂结合硅胶柱层析纯化环孢菌素A

采用大孔吸附树脂层析结合硅胶柱层析,对环孢菌素A的分离纯化进行研究,确定了最佳层析条件,建立了工业化制备环孢菌素A的工艺。大孔吸附树脂层析选用D101树脂作为吸附介质,提取液丙酮含量控制在50%,最大吸附量为35 mg/g湿树脂,洗脱剂选用丙酮;硅胶柱层析选用42~64μm硅胶作为层析介质,最优层析条件为柱床高径比10∶1,流动相配比V(石油醚)∶V(丙酮)=70∶30,流速80 mL/m in,环孢菌素A上样质量浓度100 g/L,硅胶层析平均收率为84.2%,环孢菌素A纯度可达到97%以上,整个工艺总收率为65%~70%。     

环孢菌素A结晶工艺的优化

为了确定环孢菌素A结晶工艺,对溶剂种类、反溶剂加入量、结晶温度和降温方式等因素的影响进行研究。首先通过静态结晶研究,确定了丙酮／水结晶体系和溶剂比例。在此基础上,设计了正交试验L9（34）考察动态结晶中各因素对环孢菌素A结晶收率和纯度的影响,并进一步优化了结晶降温方式。结果表明：确定环孢菌素A结晶的最佳条件为采用梯度程序降温,养晶温度为-5℃,丙酮和水的体积比为2：1,反溶剂水流加时间为0．5h,开始流加点为降温至0℃,结晶时间约为3h。经HPLC分析环孢菌素A的纯度平均值为99．15％,收率平均值为87．7％。     

基于微生物同化作用的D-丙氨酸生产工艺研究

以L-丙氨酸为唯一碳氮源,从采集的若干土壤中初筛出能够降解L-丙氨酸的菌株;再以D-丙氨酸为唯一碳氮源,复筛出降解L-丙氨酸而不降解D-丙氨酸的菌株。依据菌种对DL-丙氨酸的不对称降解活性,筛选出具有最高的L-丙氨酸降解活性的菌株,并对菌株同化L-丙氨酸的反应条件进行了研究。结果表明:编号为ALA-D82的菌株具有最高的降解L-丙氨酸的能力,经鉴定为酵母菌属。在30℃,控制pH6.0,通气比1:1(V/V)和转速900 r.min-1的条件下,L-丙氨酸降解的速度最大。在最适条件下,1500 g DL-丙氨酸分两部分添加入7 L的反应液中。反应72 h后溶液中的L-丙氨酸被完全降解,提取得到D-丙氨酸晶体,产率和光学纯度分别达到92.13%和99%。     

β-Homo天冬酰胺的合成及其副反应的研究

以α L 天冬酰胺为研究对象 ,应用Arndt Eistert反应 ,合成了 β Homo 天冬酰胺 ;同时合成了两种脱水产物β 氰基丙氨酸甲酸和 β 氰基丙氨酸重氮酮 ,对它在此反应的脱水机理进行了探讨。     

ARTP诱变选育环孢菌素A高产菌株及发酵工艺优化

采用常压室温等离子体技术(Atmospheric Room Temperature Plasma,ARTP)对雪白白僵菌FIM-1809菌株进行诱变,得到一株遗传稳定性较高的突变高产菌FIM-1809-7,其产环孢菌素A能力较初始菌株提高约39%。通过单因素和正交设计试验优化,确定最佳发酵培养基组分及培养条件为：玉米浆粉6%、可溶性淀粉12%、葡萄糖1.2%、蛋白胨2%,pH 5.6,菌种菌龄为72 h,接种量10%,装液量70 m L/500 m L,发酵时间8 d,最终突变菌株产环孢菌素A效价较出发菌种增加了约53%。结果表明,采用ARTP技术结合发酵条件优化能够有效提高雪白白僵菌产环孢菌素A的能力,为该菌株的工业化应用奠定了良好的基础。     

酶法合成中丙氨酸的检测

本文建立了丙氨酸在醋酸缓冲液中形成丙氨酸-铜配离子及其十二烷基磺酸配离子对,使其紫外无吸收的丙氨酸在230nm处于有强紫外吸收,从而对酶法合成中的丙氨酸进行定量分析．该法在0～50mg／L范围内有良好的线性关系,直线方程为A(230)=0.01712·C（n=5）C：mg／L）。相关系数为0．9994,回收率为96．8％～104．0％,且该法不受酶反应液的影响,实验结果证明该检测体系简单、实用、测定结果可靠。     

利用麦芽糖假丝酵母的不对称降解活性由d1-丙氨酸生产d-丙氨酸

为了开发一种由dl-丙氨酸生产d-丙氨酸的实用工艺,我们依据酵母对dl-丙氨酸的不对称降解的活性,筛选了107株酵母菌。麦芽糖假丝酵母JCM1504降解l-异构体比d-异构体快10倍,该菌株被用来作为去除l-异构体的生物催化剂。但是,当降解反应在高浓度dl-丙氨酸下进行时,反应混和物的pH值由于降解l-丙氨酸时释放出氨而迅速地升高,因此反应停止。这个障碍可以采用在反应过程中加入H_2SO_4控制pH6.0的办法来克服。另外,我们发现,l-异构体降解的最大速度在30℃,pH6.0高通气量(1.0vvm)和振荡(1200r/min)的情况下得到。在最适条件下,dl-丙氨酸200g/L中的l-异构体在40小时内完全降解掉,在反应混和物中留下90gd-丙氨酸。d-丙氨酸很容易从反应混和物中分离出来。这样得到的d-异构体的化学和旋光纯度分别达到99.0%和99.9%。     

D—丙氨酸对不同日龄家蝇的影响

D-丙氨酸可以明显缩短家蝇(Musca Domestica)的寿命,给不同日龄的家蝇喂养1%D-丙氨酸水溶液后,出现死亡高峰的时间不同,如1日龄为8天,16日龄为4天。这种差异可能是由于机体对氨基酸的光学异构体的识别能力随龄下降,大量D-丙氨酸与L-丙氨酸竞争,参与各种L-丙氨酸所参与的反应和蛋白合成过程,使代谢失调及反应误差率随龄增加,从而加速死亡的结果。     

The role of posttranslational modification in moonlighting glyceraldehyde-3-phosphate dehydrogenase structure and function

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a moonlighting protein exhibiting distinct activities apart from its classical role in glycolysis. Regulation of its moonlighting functions and its subcellular localization may be dependent on its posttranslational modification (PTM). The latter include its phosphorylation, which is required for its role in intermembrane trafficking, synaptic transmission and cancer survival; nitrosylation, which is required for its function in apoptosis, heme metabolism and the immune response; acetylation which is necessary for its modulation of apoptotic gene regulation; and N-acetylglucosamine modification which may induce changes in GAPDH oligomeric structure. These findings suggest a structure function relationship between GAPDH posttranslational modification and its diverse moonlighting activities.

     

Characterisation of a new species of Pythium isolated from a wheat field in northern France and its antagonism towards Botrytis cinerea causing the grey mould disease of the grapevine

A new species, Pythium bifurcatum, isolated from soil samples taken from a wheat field in Lille in northern France is described here. The oomycete occurred thrice out of 50 samples. The type specimen is F-91, which is a slow-growing saprophyte living on vegetable debris and which can be recognised by its antheridial as well as oogonial characteristics, which are different from other known species of Pythium. When grown together with Botrytis cinerea, the causal agent of the grey mould disease of the grapevine, Pythium bifurcatum shows a pronounced antagonism and suppresses its growth. Morphological features of this new species, its antagonism to B. cinerea, the sequences of the ITS region of its nuclear ribosomal DNA, and its comparison with related species are discussed in this article.     

中华蟾蜍颈动脉腺的组织学结构

2005年10月～2006年5月,用组织学技术研究中华蟾蜍颈动脉腺结构。结果表明,颈动脉腺位于外颈动脉基部,圆球形,深红色至棕褐色。组织结构显示：颈动脉腺的外壁是动脉管壁的延续,包括外膜、中膜和内膜。整个外壁厚薄不均。颈动脉腺的最大特点是中膜和内膜并不像一般血管形成环圈状,而是从不同部位向管腔突出延伸、相互连接构成大小不一、形状各异、迂回曲折的网状血管。管腔大者,管壁厚,弹性纤维、平滑肌纤维多,内膜靠腔面内皮细胞多成立方状,细胞核端位近圆形;管腔小者,管壁薄,弹性纤维、平滑肌纤维少,内皮细胞扁平、排列稀疏,胞核长梭形;一些区域管径极小,管壁极薄,成为开放的血窦,只允许一个血细胞通过。网状管壁间有密集成团的大型类上皮细胞等细胞分布。据结构推测中华蟾蜍颈动脉腺有调节血压等功能。     

Partial purification and substrate analysis of bacterially expressed HIV protease by means of monoclonal antibody

Retroviruses code for a specific protease which is essential for polyprotein precursor processing and viral infectivity. The HIV-specific protease has been predicted to be an aspartic protease which is located at the amino terminus of the pol gene. We have prepared several constructs for bacterial expression of the protease. Two of them span the whole protease region and result in its autocatalytic activation. Analysis of the dynamics of this activation indicates a two-step process which starts at the carboxy terminus and ends at the amino terminus of the protease. The activated protease is a molecule of 9 kd as evidenced by monoclonal antibody in immunoblot analysis. A construct in which the carboxy terminus of the protease is deleted results in a stable, enzymatically inactive 27-kd protein which proved useful as substrate since it contains one of the predicted cleavage sites. The stability of this protein indicates that the carboxy-terminal sequences of the protease are essential for its activity and its autocatalytic activation. The protease which is very hydrophobic was solubilized by acetone treatment and passaged over ultrogel and propylagarose columns for partial purification. It elutes as a dimer and tends to aggregate. It is inhibited by pepstatin A in agreement with its expected active site and its theoretical classification as aspartic protease. Cleavage of the gag precursor results in the mature capsid protein, p17. The protease does not, however, cleave the denatured 27-kd substrate or the denatured gag precursor. Therefore its specificity appears to be not solely sequence- but also conformation-dependent. This property needs to be taken into account for the development of protease inhibitors for therapy of AIDS.     

Male‐female differences in family size preferences among college students

Abstract

Three dimensions of isolated spousal separation largely determine its fertility impact: (1) length, (2) “completeness” (degree to which it is eliminating fecundability during fecundable months), and (3) its timing as defined by the reproductive state of the wife at its start. While it is obvious that by starting when the wife is in the fecundable state its net delaying of next conception is maximized, it is much less clear with which reproductive state its start should coincide in order to minimize its fertility impact. By means of a simple model, it is shown that for minimum impact a separation should start later in a pregnancy the longer is post‐partum anovulation, the higher is the level of intrauterine mortality, and the shorter is the period of separation. Conditions of natural fertility are assumed.     

青天葵野生变家栽的探讨——Ⅰ.生物学特性的观察

青天葵是广西特产药材,过去一直处于野生状态,药源日渐枯竭.供不应求。作者通过调查和研究,掌握了它的生态条件和生物学特性。实践证明,青天葵由野生变为人工栽培是可能的。     

Bacteriocin small of fast-growing rhizobia is chloroform soluble and is not required for effective nodulation.

Small bacteriocin is a low-molecular-weight bacteriocin which is common in fast-growing rhizobia. As its activity could not be detected in chloroform-sterilized culture supernatants (P.R. Hirsch, J. Gen. Microbiol. 113:219-228, 1979), the bacteriocin could not be purified in order to study its mechanism of action. We report here that small is soluble in chloroform, an observation which led to effective and simple (partial) purification. Other properties of small are its low molecular weight, which is estimated to be between 700 and 1,500, its resistance to proteolytic enzymes, pectinase, and lysozyme, and its heat stability at pH 5.5 but not at pH 7.0. Its bactericidal action on exponentially growing sensitive cells was not detected until 11 h after its addition. The bactericidal action was preceded by inhibition of cell division. To determine whether small activity is required for nodulation or nitrogen fixation, a transposon Tn5-induced small-negative mutant was isolated. The observation that this strain formed normal, acetylene-reducing root nodules showed that small production is not a prerequisite for the formation of effective nodules.     

Antithrombin: in control of coagulation

Antithrombin is a serine proteinase inhibitor (serpin) which controls the process of coagulation. It has a well defined structure, consisting of three beta-sheets, nine alpha-helices and a reactive centre loop (RCL). The RCL contains the reactive centre which harbours a bait sequence for target proteases; cleavage results in inhibition by a unique mechanism. The inhibitory activity of antithrombin is controlled by its interaction with the co-factor, heparin, which accelerates its interaction with target proteases. This ensures that heparin and its newer derivatives, such as heparin pentasaccharide, are the mainstay therapeutics for control of thrombosis or inappropriate clotting. The clinical importance of antithrombin is manifested by its clear association with thrombosis when deficiency states occur.     

Mechanism and stereospecificity of the parabactin-mediated iron-transport system in Paracoccus denitrificans

The mechanism by which Paracoccus denitrificans utilizes parabactin in its iron-transport apparatus is examined. The cellular uptake of 55Fe-parabactin, its enantiomer, as well as a large number of its homologues and analogues are measured. Furthermore, the ability of these catecholamide ligands to stimulate microbial growth is also determined. The results of these studies point out several structural boundary conditions which the microorganism sets when utilizing iron chelates.     

城口荨麻,重庆荨麻科一新种

描述了自重庆市发现的荨麻科荨麻属一新种,城口荨麻,此种与异株荨麻有亲缘关系,区别特征为此种的茎被少数刺毛,叶片多为心形,雄、雌花序均不分枝,瘦果在中央稍凹陷。