Localisation of substance P-like immunoreactivity in the ciliary ganglia of monkey (Macaca fascicularis) and cat: a light- and electron-microscopic study

The present study describes substance P-like immunoreactivity in the ciliary ganglia of monkey (Macaca fascicularis) and cat. About 60% of neurons in the monkey ciliary ganglion and 40% in the cat ciliary ganglion were substance P-like immunoreactive, ranging from faint to moderate staining. Substance P-like immunoreactivity was located in cell bodies, dendritic profiles and axons. In the monkey, substance P-like immunoreactive pericellular arborisations were associated with about 0.5%–3% of the ganglion cells, which were either negatively, faintly or moderately stained. An electron-microscopic study demonstrated the presence of either substance P-like immunoreactive positive or negative axon terminals synapsing or closely associated with positive dendritic profiles in both the monkey and cat ciliary ganglia. The results suggest that substance P plays an important role in the ciliary ganglion, perhaps as a modulator or transmitter.     

含金地质体表生带中主要微生物类群的研究

通过对16个含金矿区主要微生物类群的分布、生态结构的研究,发现不同类型的金矿具有不同的微生物类群.金矿矿石与金矿酸水中的自养硫细菌和异养菌分布具有明显差异,酸性水中含自养硫细菌较多,含异养菌极少;金矿氧化带中的异氧菌一般多于自氧硫细菌,其优势菌以芽孢杆菌属中蜡状芽孢杆菌为主.研究结果揭示了金矿中存在的主要微生物类群,对阐明表生金的形成、成岩和蚀变作用具有一定的意义,对发掘生物湿法冶金菌种资源具有重要作用.     

The effect of ferredoxin(BED) overexpression on benzene dioxygenase activity in Pseudomonas putida ML2.

The benzene dioxygenase from Pseudomonas putida ML2 is a multicomponent complex comprising a flavoprotein reductase, a ferredoxin, and a terminal iron-sulfur protein (ISP). The catalytic activity of the isolated complex shows a nonlinear relationship with protein concentration in cell extracts, with the limiting factor for activity in vitro being ferredoxin(BED). The relative levels of the three components were analyzed by using 125I-labelled antibodies, and the functional molar ratio of ISP(BED), ferredoxin(BED), and reductase(BED) was shown to be 1:0.9:0.8, respectively. The concentration of ferredoxin(BED) was confirmed by quantitative electron paramagnetic resonance spectroscopy of the 2Fe-2S centers in ferredoxin(BED) and ISP(BED) of whole cells. These results demonstrate that the ferredoxin(BED) component is a limiting factor in dioxygenase activity in vitro. To determine if it is a limiting factor in vivo, a plasmid (pJRM606) overproducing ferredoxin(BED) was introduced into P. putida ML2. The benzene dioxygenase activity of this strain, measured in cell extracts, was fivefold greater than in the wild type, and the activity was linear with protein concentration in cell extracts above 2 mg/ml. Western blotting (immunoblotting) and electron paramagnetic resonance spectroscopic analysis confirmed an elevated level of ferredoxin(BED) protein and active redox centers in the recombinant strain. However, in these cells, the increased level of ferredoxin(BED) had no effect on the overall rate of benzene oxidation by whole cells. Thus, we conclude that ferredoxin(BED) is not limiting at the high intracellular concentration (0.48 mM) found in cells.     

Stomatal sensitivity to abscisic acid following water deficit stress

Short-and medium-term stresses (1 and 24 h, respectively) wereapplied to detached leaves of Commelina communis L., resultingin both cases in a final leaf cell water potential (     

Nanometric design of extraordinary hydrophobic-induced pKa shifts for aspartic acid: Relevance to protein mechanisms

Commonly a key element enabling proteins to function is an amino acid residue or residues with functional side chains having shifted pKa values. This article reports the results on a set of protein-based polymers (model proteins) that exhibit hydrophobic folding and assembly transitions, and that have been designed for the purpose of achieving large hydrophobic-induced pKa shifts by selectively replacing Val residues by Phe residues. The high molecular weight polypentapeptides, actually poly (tricosapeptides) with six varied pentamers in fixed sequence, were designed and synthesized to have the same amino acid compositions but different proximities between a single aspartic acid residue and 5 Phe residues per 30 residues. With the 5 Phe residues distal from the Asp residue, the observed pKa shift was 2.9 when compared to the Val-containing reference. With the 5 Phe residues within 1 nm of the Asp residue, the pKa shift was 6.2. This represents a free energy of interaction of 8 kcal/moles. To our knowledge, this is the largest pKa shift documented for an Asp residue in a polypeptide– or protein–water system. Data are reviewed that do not support the usual electrostatic arguments for pKa shifts of charge–charge repulsion and/or unfavorable ion self-energies arising from displacement of water by hydrophobic moieties, but rather the data are interpreted to indicate the presence of an apolar–polar repulsive free energy of hydration, which results from a potentially highly cooperative competition between apolar and polar species for hydration. © 1994 John Wiley & Sons, Inc.     

An enzyme-immobilized microplate for determination of linamarin for large number of samples

Summary An enzyme-immobilized microplate for determination of linamarin was prepared by covalently linking cassava leaf linamarase to the microplate. For linamarin determination, cassava roots were homogenised in 0.1 Mo-phosphoric acid and the filtrate adjusted to pH 6 with NaOH prior to adding into the wells. The cyanide released was then determined spectrophotometrically. One nmol linamarin can be detected. The microplate method is suitable for analysis of large number of samples and is useful for screening purposes.     

Introduction of oxygen into the alkyl chain of N-decyl-dNM decreases lipophilicity and results in increased retention of glucose residues on N-linked oligosaccharides

N-Alkylation of the -glucosidase inhibitor 1-deoxynojirimycin(dNM) dramatically increases its inhibitory potency (Tan etal., J. Biol. Chem., 266, 14504–14510, 1991). However,the possibility of extending the alkyl chain to N-decyl-dNMis limited by an increase of detergent-like (amphiphilic) propertiesof long-chain alkylated dNM derivatives. Substitution of methylenegroups in the N-decyl chain by oxygen reduced the amphiphilicityof N-decyl-dNM derivatives, while retaining their superior inhibitoryproperties. In intact HepG2 cells, the compound N-7-oxadecyl-dNMwas found to result in the most pronounced retention of glucoseresidues on N-linked glycans. Permeabilization of the plasmamembrane with the bacterial toxin Streptolysin O improves theinhibitory properties of the derivatives N-3,6,9-trioxadecyl-,N-7,10,13-trioxatetradecyl-, N-3-oxadecyl- and N-7-oxadecyl-dNM,but not those of dNM. These observations suggest differencesin the mode of entry of the oxygen-substituted dNM derivativesin comparison with dNM. We observed that the dNM derivativeN-3,6,9-trioxadecyl-dNM, devoid of inhibitory activity in intactcells, was inhibitory in Streptolysh O-permeabilized cells.Thus, the permeability barriers posed by plasma membrane andendoplasmic reticulum membrane are not equivalent. The use ofa permeabilized cell system thus allows the elaboration of inhibitoryprinciples for novel bioactive compounds where study of theisolated enzymes may not be possible, and where intact cellsare not a suitable target due to permeability barriers. -glucosidase inhibition N-linked glycosylation oxygen-substituted N-decyl-dNM derivatives permeabilized cells     

番木瓜环斑病毒复制酶(亚基)基因的克隆、序列分析及其植物表达载体的构建

近二年报道,在TMV^⑴、PVX^⑵、PEBV^⑶和CMV^⑷等病毒中,利用病毒编码的复制酶(亚基)基因或相关cDNA片段转化烟草,工程植株获得绝对或极高的病毒抗性。大量研究认为：马铃薯Y病毒组的核内含体大分子量蛋白(Nib)是依赖于RNA的RNA复制酶(或核心亚基),Nlb与上述病毒的复制酶有广泛的同源保守区^⑸。因此,Nib基因的克隆,不但在植物抗病基因工程方面,而且在马铃薯Y病毒组基因组的复制研究方面．均有重要的意义。本文以马铃薯Y病毒组的重要成员番木瓜环斑病毒的华南强株系(PRSV—sM)为材料,克隆了PRSV的Nib基因,并完成其全序列测定和植物表达载体的构建,为探索马铃薯Y病毒组复制酶可能介导的抗病性打下了基础。     

卡拉胶固定粘质赛氏菌产碱性蛋白酶的研究

将粘质赛氏菌(Seratia marcescens)包埋于卡拉胶中,发现2．5％的卡拉胶适于固定该菌产碱性蛋白酶。固定化细胞在其较适宜产酶培养基中发酵,酶活力一般可达400u/ml,在卡拉胶中添加3％玉米粉和1%豆饼粉或2％砂子制备固定化细胞,其产酶能力分别提高了25%和23．9％;固定化细胞颗粒越小,其产酶能力越高。采用摇瓶半连续发酵。其产酶半衰期为14次(24小时为一个周期);而用环流器进行半连续发酵,其产酶半衰期为52次(12小时为一个周期),产酶效率分别比游离细胞摇瓶发酵的产酶效率高11．8％和45．07％,而环流器半连续发酵的产酶效率比摇瓶半连续发酵高29．7％。