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Chinese Science Bulletin 2000年第1期45卷 57-60页

作者： Hui Yu Bin Kang Jie Wei Liangbi Li Tingyun Kuang Chinese Acad Sci Inst Bot Photosynth Res Ctr Beijing 100093 Peoples R China

The 33 ku protein, prepared from NaCI-treated PSII particles, has shown an single band by SDS-PAGE. After being dialyzed against the low-osmotic medium at 4癈, it has been found that the 33 ku protein degraded into several small fragments. This result suggests that the preparations of 33 ku protein probably contain some latent proteinases. it has also been found, by the 2-D electrophoresis and IEF, that the preparations of 33 ku protein not dialyzed against the low-osmotic medium contain several polypeptides with nearly the same molecular weight but not the same isoelectric point as the 33 ku protein.

关键词： 33 ku protein proteinase isoelectric point isoelectric focusing 2-D (2-dimensional) electrophoresis.

Bio-Control of Alternaria alternataduring Banana Storage by Purified AFP Using isoelectric Focusing Technique

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Food and Nutrition Sciences 2014年第15期5卷 1482-1495页

作者： Hassan Barakat Department of Food Science Faculty of Agriculture Benha University 13736 Moshtohor Kaliuobia Egypt

Interestingly, antifungal protein AFP was purified from Aspergillus giganteus supernatants with modified isoelectric focusing procedure after adaptation of the secretion conditions. Subsequently, the antifungal activity as well as the mode of action against Alternaria alternata was tested in vitro. Moreover, different concentrations of AFP were applied to banana fruits for 15 days at 20°C in vivo. Obtained results illustrated that A. giganteus was able to secrete about 39.78 ± 2.39 mg AFP·l-1 Olson medium. The employed ammonium sulfate (AS 75%) precipitation procedure followed by dialysis steps yielded about 16 - 22 mg AFP·l-1 culture supernatant with general mean of 18.67 ± 1.98 mg·l-1. The lost amount of AFP during purification using AS and 3KDa cut-off dialysis membrane is about 50% thus, purification procedure must be further improved. Indeed, concluded results from MIC and hyphal extension inhibition test noticed that AFP was efficiently affected by either growth or hyphae form of A. alternata in vitro. The MIC of AFP against A. alternata was 2μg·ml-1. However,short, thick and highly septated hyphae with damaged constricted apical regions extruding from condensed mycelium aggregates in treated hyphae compared to the untreated culture was remarkably shown. The mode of action of in vitro experiment manifested that AFP was effective to act the fungal cell and permeabilize the cell membrane of A. alternata. Furthermore, the in vivo experiment showed that AFP could reduce post-harvest decay on banana caused by A. alternata. AFP at concentration of 15 and 25 μg·ml-1 exhibit Alternaria decayed reduction by 45.45% and 77.27%, respectively. While no Alternaria decayed area was observed when 50 μg·ml-1 was applied during the storage time. Quantification of DNA by species-specific PCR could exude a positive correlation between the DNA amount and decayed area. In conclusion, AFP can be efficiently used as a bio-preservative agent during storage and handling of banana

关键词： Antifungal Protein (AFP) Alternaria alternata isoelectric Focusing Antifungal Activity

Sensitivity and Specificity Determinations with isoelectric Focusing Fractions of Blastomyces dermatitidisfor Antibody Detection in Serum Specimens from Infected Dogs

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Open Journal of Veterinary Medicine 2012年第4期2卷 237-241页

作者： Joshua C. Wright Terrick E. Harrild Gene M. Scalarone Department of Biological Sciences Idaho State University Pocatello USA

Blastomycosis and histoplasmosis manifest as lung and systemic fungal infections in mammals caused by Histoplasma capsulatum, and Blastomyces dermatitidis. These infections exhibit cross reactivity of antibodies which makes a correct diagnosis potentially elusive. The purpose of this study was to gain an understanding of which isoelectric focusing fractions (RotoforTM) of B. dermatitidis were reactive or cross reactive with serum specimens from dogs infected with B. dermatitidis, H. capsulatum, and Cryptococcus neoformans. Three serum specimens from dogs that were infected with B. dermatitidis, two dogs infected with H. capsulatum, and one dog infected with C. neoformans were assayed against the 20 B. dermatitidis RotoforTM fractions. Reactivity was determined using the indirect enzyme linked immunoassay (ELISA). Reactivity with B. dermatitidis was found predominantly in the protein fractions 1 - 6, and cross reactivity with H. capsulatum, and C. neoformans sera was found within the B. dermatitidis protein fractions 15 - 19.

关键词： isoelectric Focusing ELISA Blastomycosis Lysate Antigen Antibody Detection

Preparation of polyacrylamide based monolith with immobilized pH gradient and its application for protein analysis

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Science China Chemistry 2007年第4期50卷 526-529页

作者： ZHU GuiJie ZHANG WeiBing ZHANG LiHua LIANG Zhen ZHANG YuKui National Chromatographic R.&A.Center Dalian Institute of Chemical PhysicsChinese Academy of SciencesDalian 116023China Graduate University of Chinese Academy of Sciences Beijing 100039China

Monolithic materials were prepared in capillaries by in situ polymerization of acrylamide, glycidyl methacrylate and N,N′-memylenebisacrylamid in the presence of trinary porogens, including 1,4-butanediol, dodecanol and dimethyl sulphoxide. With Ampholine immobilized on the monolith by chemical bonding according to their p/s, the monolithic immobilized pH gradient (M-IPG) was prepared, and applied to the separation of four standard proteins. Compared with polyacrylate based M-IPG, the hydrophilicity of the new material was improved. It could not only avoid the adsorption of proteins, but also make the synthesized procedure simple, which showed great potential in the analysis of proteins.

关键词： capillary isoelectric focusing monolith immobilized pH gradient proteins

Glutathione S-Transferase Isoenzymes as Marker in Preneoplasia of Rat Chemical Hepatocarcinogenesis

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Chinese Science Bulletin 1993年第7期38卷 602-607页

作者：左瑾方福德丁濂谢大英 Institute of Basic Medical Sciences Chinese Academy of Medical Sciences Beijing 100005 PRC Institute of Occupational Medicine Chinese Academy of Preventive Medicine Beijing 100050 PRC

It has been generally accepted that about 85％ of cancers are caused by chemical factors and the chemically induced carcinogenic process can be divided into two or three stages, i. e. initiation and promotion, or together with further progression. The initiation is usually considered to be an irreversible process, but it can be blocked to further progress to cancer. For this reason, searching of objective markers for identifying initiated cells has played an important role in defining the preneoplastic lesion. So far, a large number of studies have been carried out on this object, including biochemistry, histopathology, histochemistry, immunohistochemistry, etc., and nearly thirty histochemical and other phenotypic changes have been found associated with the putative preneoplastic liver

关键词： GST-isoenzyme chemicai hepatocarcinogenesis isoelectric focusing marker enzyme.

On the Distribution of Acidic Proline-rich Proteins Phenotypes in Chinese Han Population in Liaoning Area

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中国医科大学学报 1991年第S2期20卷 33-36页

作者：庞灏贾静涛胡忠国 Department of Forensic Serology Faculty of Forensic Medicine China Medical University Shenyang China

The distribution of salivary acidic proline-rich proteins pheno-types was investigated by using the ultra-thin PAGIEF technique in 258 ChineseHan populations in Liaoning area. The gene frequencies were as follows: Pr^10.8101, Pr^20.1899; Db^+0.0416, Db^-0.9584; Pa^+0.1717, Pa^-0.8283; PIF^+0.6647, PIF^-0.3353. The observed numbers of the phenotypes are in good a-greement with the expected numbers under the Hardy-Weinberg *** gene frequencies among the Chinese and other populations are compared.

关键词： acidic proline-rich proteins polyacrylamide gel isoelectric focusing gene frequency

Responses of bioenergy sorghum cell wall metabolism to agronomic practices

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Advances in Biological Chemistry 2014年第1期4卷 67-78页

作者： Jason P. Wight Frank M. Hons Godson O. Osuji Biochemistry Laboratory Cooperative Agricultural Research Center Prairie View A&M University Prairie View USA Department of Soil and Crop Sciences Texas A&M University College Station USA

Maximum lignocellulose yield of biomass sorghum [Sorghum bicolor L. (Moench.)] is hampered by complex biological phenomena related to rotation, nitrogen (N) fertilization, soil tillage, and excessive biomass removal. The biochemical basis of the effects of agronomic practices on sorghum production was studied by the enzymology of the active peroxidase (EC 1.***.1.7) isoenzymes that synthesize lignin. All studied practices altered the peroxidase pI values. Control sorghum without rotation and without N fertilization had the most inhibited peroxidase with very low maximum velocity (Vmax) value (3.10 mmol·min﹣1), and very low lignin (857 kg·ha﹣1) yield, which could decrease soil organic carbon possibly leading to adverse changes in soil chemistry. Corn-sorghum rotations with and without N fertilization increased the Vmax values of peroxidase and lignin and cellulose yields. Rotated sorghum subjected to 50% residue return (the percentage of crop residue was returned to the plot immediately after grinding at harvest) and 280 kg·ha﹣1 N fertilization possessed very active peroxidase (Vmax value 66.4 mmole·min﹣1) and the highest lignin (1387 kg·ha﹣1) yield. The 25% residue return rate without N fertilization induced high lignin (1125 kg·ha﹣1) and cellulose (11,961 kg·ha﹣1) but the 25% residue return rate with 280 kg·ha﹣1 N fertilization induced lower lignin (1046 kg·ha﹣1) yield. Continuously cropped sorghum treated with 336 kg·N·ha﹣1 produced active peroxidase that shared competitive inhibition relationship with the peroxidase of the 84 kg·N·ha﹣1 treatment. Ridge tillage combined with 280 kg·ha﹣1 N fertilization under continuous sorghum resulted in inhibited peroxidase possessing low Vmax value (13.0 μmole·min﹣1). Changing to conventional tillage combined with 280 kg·ha﹣1 N fertilization relieved the inhibition and increased the Vmax value to 23.7 mmol·min﹣1. These biological anomalies of sorghum cell wall related to agronomic practices originated from doubly inhibited sorgh

关键词： Peroxidase Enzymology Free Solution isoelectric Focusing Lignin and Cellulose Biochemical Mechanism of Crop Rotation

Pseudomonas aeruginosaBUP2—A Novel Strain Isolated from Malabari Goat Produces Type 2 Pyoverdine

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Advances in Bioscience and Biotechnology 2014年第11期5卷 874-885页

作者： Kizhakkepowathial Nair Unni Prakasan Priji Valerie A. Geoffroy Mukesh Doble Sailas Benjamin Enzyme Technology Laboratory Biotechnology Division Department of Botany University of Calicut Malappuram India Laboratoire de Microbiologie et de Génétique CNRS UPRES-A 7010 Université Louis-Pasteur Strasbourg France Department of Biotechnology Indian Institute of Technology Madras Chennai India

This study focuses on the isolation and characterization of a novel strain of siderophore producing bacterium, i.e., Pseudomonas aeruginosa BUP2 (Pa BUP2) from the rumen of Malabari goat, coupled with qualitative and quantitative analyses of the siderophore produced by it. Pa BUP2—a facultative anaerobe was tuned to be an aerobe by repeatedly growing in Benjamin flask. The new isolate was grown in a specially designed semi-synthetic medium, designated as BUP medium, and the yellowish-green pigment produced was identified as a typical siderophore by spectrophotometry, Chromazurol-S assay, thin layer chromatography and isolectric focusing (IEF). The characteristic orange fluorescence upon UV irradiation on chromatogram and absorption maximum at λ404 confirmed that the characteristic siderophore produced by Pa BUP2 was a typical pyoverdine (PVD). This PVD was further categorized under type 2 by comparing its profile on the IEF gel with that of the representative strains of each PVD types, viz., Pa O1, Pa ATCC 27853 and Pa6. Moreover, the type 2 PVD was purified by XAD-4 Amberlite column chromatography and quantified;maximum yield (11.17 mg/ml) was observed on day 4 of incubation (37°C). Thus, it was confirmed that the bacterium isolated from the rumen content of Malabari goat is a novel strain of Pa capable of producing large quantity of PVD type 2 in specially designed BUP medium under aerobic condition, and that its clinical and industrial implications remain elusive.

关键词： Pseudomonas aeruginosa BUP2 Siderophore Pyoverdine Type 2 isoelectric Focusing

Interaction of Coal Humic Acids with Fungal Laccase

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Advances in Microbiology 2013年第2期3卷 145-153页

作者： Natalia A. Kulikova Valentina N. Davidchik Eugenia A. Tsvetkova Olga V. Koroleva Department of Soil Science M.V. Lomonosov Moscow State University Moscow Russia

Humic acids (HA) are one of the main environmental factors controlling the fate and behavior of the compounds released into the environment. In particular, they are universally considered of great importance in determining soil extracellular enzyme activity and stability via association with essential soil enzymes. The objective of this study was to investigate the interaction of coal HA with an extracellular multicopper oxidase laccase (EC 1.***.3.2) that catalyze the oxidation of a wide range of reducing substances in the environment. Using size-exclusion chromatography analysis and monitoring laccase activity, the formation of a stable and an enzymatically active complex between HA and laccase was shown. Basing the data obtained by isoelectric focusing of HA-laccase complex, non-covalent character of laccase association with HA was considered and binding of laccase to HA by weak dispersive forces such as van der Waals, hydrophobic, π-π, CH-π and others was hypothesized.

关键词： Humic Acids Laccase Activity Size-Exclusion Chromatography Complexes isoelectric Focusing

Studies on the Thiol Components of Isolated Nuclei

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Journal of Analytical Sciences, Methods and Instrumentation 2020年第1期10卷 36-42页

作者： Michael Gronow Cambridge Cancer Research Fund Laboratory 7 The Maltings Cottenham Cambridge England UK

The thiol components of the nonhistone proteins prepared from isolated nuclei from rat liver, regenerating liver and hepatoma 223 cells have been investigated after reaction with radio labelled N-ethylmaleimide and 5-5’-dithiobis-(2-nitrobenzoic acid) (DTNB). The labelled adducts formed were examined by isoelectric focusing analysis in polyacrylamide gel and the distribution of the radiolabels within sliced portions of the gels determined. In the case of the 14C labelled NEM adduct the label was found to be spread amongst numerous protein components within the gel however, in the case of the 35S labelled DTNB adducts, only a small proportion of the label was found in the protein material which was retained in the acidic isoelectric point (pI) region of the gel. The bulk of the 35S labelled adduct (56% - 60%) was found to have migrated into the anode solution (10 mM phosphoric acid). This could be adsorbed onto a hydrophobic resin (XAD2) resin and eluted with methanol. Gel filtration chromatographic analysis of this adduct material on BioGel P2, (exclusion limit 1500 daltons) showed low molecular weight components to be present. Slightly different patterns were obtained for these nuclei, each containing several 35S components with molecular weights greater than the Ellman reagent itself. These 35S labelled thiol components did not contain any protein, peptide or amino acid components indicating strongly that a novel species of thiols could be present in these nuclei bound within the non-histone protein matrices.

关键词： Thiols Isolated Cell Nuclei Radiolabelled Thiol Adducts isoelectric Focusing Analysis