摘要：
A proteomic approach was employed to investigate the cold stress-responsive proteins in trifoliate orange (Poncirus trifoliata (L.) Raf.), which is a well-known cold tolerant citrus relative and widely used as rootstock in China. Two-year-old potted seedlings were exposed to freezing temperature (-6A degrees C) for 50 min (nonlethal) and 80 min (lethal), and the total proteins were isolated from leaves of the treated plants. Nine differentially accumulated proteins over 2-fold changes in abundance were identified by two-dimensional gel electrophoresis and mass spectrometry. Among these proteins, a resistance protein induced by the nonlethal cold treatment (protein spot #2 from P. trifoliata) was selected as target sequence for degenerated primer design. By using the designed primers, a PCR product of about 700 bp size was amplified from P. trifoliata genomic DNA, which was further cloned and sequenced. A nucleotide sequence of 676 bp was obtained and named Ptcorp. Blast retrieval showed that Ptcorp shared 88% homology with an EST of cold acclimated Bluecrop (Vaccinium corymbosum) library (Accession number: CF811080), indicating that Ptcorp had association with cold acclimation. Semiquantitative RT-PCR analysis demonstrated that Ptcorp gene was up-regulated by cold stress which was consistent with the former result of protein expression profile. As the resistance protein (NBS-LRR disease resistance protein family) gene was up-regulated by cold stress in trifoliate orange and satsuma mandarin, it may imply that NBS-LRR genes might be associated with cold resistance in citrus.
摘要：
The study of DNA-binding proteins is crucial in understanding gene regulatory networks. We developed a new method for the enrichment of DNA-binding proteins based on the variability of DNA-protein complexes’ solubility in different ionic strength solutions. 0.14M sodium chloride was determined as the most efficient extraction concentration to precipitate DNA-binding proteins. SDS-PAGE analysis revealed that some high-abundance proteins were removed effectively and at the same time DNA-binding proteins were isolated in this simple process. Twenty kinds of proteins were identified in the acquired sample by 1-D gel-LC-MS/MS. Furthermore, computerized analysis of MS data showed that quite a number of unmatched peptides have the classic structure of leucine zipper or zinc finger, which were symbolic elements of transcription factors. These results suggested that this new method can acquire DNA-binding proteins effectively and allow improvement in the isolation of high-quality DNA-binding proteins.
摘要：
A systematic strategy was developed for the proteomic analysis of wheat chloroplast protein complexes. First, comprehensive centrifugation methods were utilized for the exhaustive isolation of thylakoid, envelope, and stromal fractions. Second, 1% n-dodecyl-β-D-maltoside was selected from a series of detergents as the optimal detergent to dissolve protein complexes effectively from membranes. Then, blue native polyacrylamide gel electrophoresis (BN-PAGE) and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) were improved to separate and analyze the protein complexes. By this systematic strategy, envelopes, thylakoids, and stromata were enriched effectively from chloroplasts in the same process, and more than 18 complexes were obtained simultaneously by BN-PAGE. Finally, thylakoid protein complexes were further analyzed by BN/SDS-PAGE, and nine complex bands and 40 protein spots were observed on BN-PAGE and SDS-PAGE respectively. Our results indicate that this new strategy can be used efficiently to analyze the proteome of chloroplast protein complexes and can be applied conveniently to the analysis of other subcellular protein complexes.
期刊：
Journal of Medicinal Plants Research,2011年5(11):2192-2200 ISSN：1996-0875
通讯作者：
Liang, Yizeng
作者机构：
[Liang, Yizeng; Tang, Zhonghai] Cent S Univ, Coll Chem & Chem Engn, Res Ctr Modernizat Chinese Tradit & Herbal Drug M, Changsha 410083, Hunan, Peoples R China.;[Shi, Guorong; Peng, Guoping; Zhou, Min; Rao, Liqun] Hunan Agr Univ, Coll Biosci & Biotechnol, Changsha 410128, Hunan, Peoples R China.
通讯机构：
[Liang, Yizeng] C;Cent S Univ, Coll Chem & Chem Engn, Res Ctr Modernizat Chinese Tradit & Herbal Drug M, Changsha 410083, Hunan, Peoples R China.
摘要：
This study examined the effects of endophytic fungus and its elicitors on cell status and alkaloid synthesis in cell suspension cultures of Catharanthus roseus. Endophytic fungus and its elicitors were inoculated in cell suspension cultures of C. roseus respectively. The biochemical change of suspension cells was then tested. The results showed when the pH rises, the concentration of MDA increases as the activities of antioxidative enzymes (peroxidase and catalase) rises. The critical enzymes of alkaloid synthesis include phenylalanine ammonialyase and tryptophan decarboxylase in suspension cells were also found to increase, along with the alkaloid yield rises. The alkaloid yields reached 770.36 693.76 and 693.76 mu g/gFW in the co-culture and elicitor-induced groups, which are 48 and 32% higher than the control group, respectively.
摘要：
Two-year-old seedlings of trifoliate orange (Poncirus trifoliata (L.) Raf.) and lemon (Citrus limonia Osbeck.) were sprayed with 0.1 mmol.L-1 abscisic acid (ABA) and then their leaves were collected randomly to expose to freezing stress. Specifically, these detached leaves were treated at 0, -3, -6, -9 and -12°C for 1 h respectively.The concentration of malondialdehyde (MDA), percentage of electrolyte leakage (EL) and the activities of several antioxidant enzymes (superoxide dismutase, catalase and peroxidase) were determined spectrophotometrically. The results show that the percentage of electrolyte leakage in trifoliate orange was lower than that in lemon, while the MDA concentration and the antioxidant activities of antioxidant enzymes in trifoliate orange were higher than those in lemon. Pretreatment with the abscisic acid can significantly reduce the membrane damage caused by freezing stress. However, the lipid peroxidation damage caused by low temperature and the activities of antioxidant enzymes were irregularly influenced by ABA pretreatment. We were thus able to infer that exogenous ABA can enhance the cold tolerance of citrus by a process rather than by activating antioxidant enzymes.
Abbreviation
ABA, Abscisic acid; POD, peroxidase; CAT, catalase; SOD, superoxide dismutase; EL, electrolyte leakage; MDA, malondialdehyde; ROS, reactive oxygen species.
摘要：
Five flavones possessing one to four phenolic groups were fully phosphorylated efficiently and the obtained compounds showed excellent pancreatic cholesterol esterase (CEase) inhibitory activities with IC(50) in the nanomolar range, which were much more potent than their parent compounds. The inhibition mechanism and kinetic characterization studies indicate that they are irreversible competitive inhibitors.
