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91.
Expressionofmilkproteingenesisinvolvedinahugenetworkofregulatorycircuitswhicharelinkedtotheintactdevelopingmammarygland,andhomeostasisduringpuberty,pregnancy,lactationandinvolution.Analysisofputativeregulatoryelementsandhybridgeneintissueculturesystems… 相似文献
92.
Cabantous S Pédelacq JD Mark BL Naranjo C Terwilliger TC Waldo GS 《Journal of structural and functional genomics》2005,6(2-3):113-119
We have improved our green fluorescent protein (GFP) folding reporter technology [Waldo et al., (1999) Nat. Biotechnol. 17, 691–695] to evolve recalcitrant proteins from Mycobacterium tuberculosis. The target protein is inserted into the scaffolding of the GFP, eliminating false-positive artifacts caused by expression
of truncated protein variants from internal cryptic ribosome binding sites in the target RNA. In parallel, we have developed
a new quantitative fluorescent protein tagging and detection system based on micro-domains of GFP. This split-GFP system,
which works both in vivo and in vitro, is amenable to high-throughput assays of protein expression and solubility [Cabantous et al., (2005) Nat. Biotechnol. 23, 102–107]. Together, the GFP folding reporter and split-GFP technologies offer a comprehensive system for manipulating and
improving protein folding and solubility. 相似文献
93.
The rat ErbB2 tyrosine kinase receptor produced in plants is immunogenic in mice and confers protective immunity against ErbB2+ mammary cancer 下载免费PDF全文
Slavica Matić Elena Quaglino Lucia Arata Federica Riccardo Mattia Pegoraro Marta Vallino Federica Cavallo Emanuela Noris 《Plant biotechnology journal》2016,14(1):153-159
The rat ErbB2 (rErbB2) protein is a 185‐kDa glycoprotein belonging to the epidermal growth factor‐related proteins (ErbB) of receptor tyrosine kinases. Overexpression and mutations of ErbB proteins lead to several malignancies including breast, lung, pancreatic, bladder and ovary carcinomas. ErbB2 is immunogenic and is an ideal candidate for cancer immunotherapy. We investigated the possibility of expressing the extracellular (EC) domain of rErbB2 (653 amino acids, aa) in Nicotiana benthamiana plants, testing the influence of the 23 aa transmembrane (TM) sequence on protein accumulation. Synthetic variants of the rErbB2 gene portion encoding the EC domain, optimized with a human codon usage and either linked to the full TM domain (rErbB2_TM, 676 aa), to a portion of it (rErbB2‐pTM, 662 aa), or deprived of it (rErbB2_noTM, 653 aa) were cloned in the pEAQ‐HT expression vector as 6X His tag fusions. All rErbB2 variants (72–74.5 kDa) were transiently expressed, but the TM was detrimental for rErbB2 EC accumulation. rERbB2_noTM was the most expressed protein; it was solubilized and purified with Nickel affinity resin. When crude soluble extracts expressing rErbB2_noTM were administered to BALB/c mice, specific rErbB2 immune responses were triggered. A potent antitumour activity was induced when vaccinated mice were challenged with syngeneic transplantable ErbB2+ mammary carcinoma cells. To our knowledge, this is the first report of expression of rErbB2 in plants and of its efficacy in inducing a protective antitumour immune response, opening interesting perspectives for further immunological testing. 相似文献
94.
95.
Chitosan functional properties 总被引:7,自引:0,他引:7
Chitosan is a partially deacetylated polymer of N-acetyl glucosamine. It is essentially a natural, water-soluble, derivative
of cellulose with unique properties. Chitosan is usually prepared from chitin (2 acetamido-2-deoxy β-1,4-D-glucan) and chitin
has been found in a wide range of natural sources (crustaceans, fungi, insects, annelids, molluscs, coelenterata etc.) However
chitosan is only manufactured from crustaceans (crab and crayfish) primarily because a large amount of the crustacean exoskeleton
is available as a by product of food processing. Squid pens (a waste byproduct of New Zealand squid processing) are a novel,
renewable source of chitin and chitosan. Squid pens are currently regarded as waste and so the raw material is relatively
cheap. This study was intended to assess the functional properties of squid pen chitosan. Chitosan was extracted from squid
pens and assessed for composition, rheology, flocculation, film formation and antimicrobial properties. Crustacean chitosans
were also assessed for comparison. Squid chitosan was colourless, had a low ash content and had significantly improved thickening
and suspending properties. The flocculation capacity of squid chitosan was low in comparison with the crustacean sourced chitosans.
However it should be possible to increase the flocculation capacity of squid pen chitosan by decreasing the degree of acetylation.
Films made with squid chitosan were more elastic than crustacean chitosan with improved functional properties. This high quality
chitosan could prove particularly suitable for medical/analytical applications.
This revised version was published online in November 2006 with corrections to the Cover Date. 相似文献
96.
Kees W. Rodenburg Marcel J. A. de Groot Rob A. Schilperoort Paul J. J. Hooykaas 《Plant molecular biology》1989,13(6):711-719
In relation to the question which DNA form (single- or double-stranded) is transferred by Agrobacterium tumefaciens to plant cells, we studied the behaviour of single-stranded DNA, as compared to double-stranded DNA, when it is introduced into plant protoplasts by electroporation. To this end, we cloned a construct with a plant NPTII gene as well as a CAT gene in the M13 vectors tg130 and tg131. We found that both complementary single-stranded molecules gave rise to substantial CAT activity in plant protoplasts, suggesting that single-stranded DNA is converted into double-stranded DNA by the plant cell replication machinery. Unexpectedly, we found that single-stranded DNA leads to a 3–10 fold higher frequency of stable transformation (selection for kanamycin resistance) than double-stranded DNA. These results indicate that the use of single-stranded DNA might be considered in experiments in which optimal transformation frequencies are needed, e.g. with protoplasts form recalcitrant plant species.Abbreviations ss
single-stranded
- ds
double-stranded
- CAT
chloramphenicol acetyl transferase
- NPTII
neomycin phosphotransferase II
- RT
room temperature 相似文献
97.
Vlasie MD Fernández-Busnadiego R Prudêncio M Ubbink M 《Journal of molecular biology》2008,375(5):1405-1415
Copper-containing nitrite reductase is able to catalyze the reduction of nitrite with a turnover rate of several hundreds per second. Electrons for the reaction are donated by the electron transfer protein pseudoazurin. The process of protein complex formation, electron transfer and dissociation must occur on the millisecond timescale to enable the fast turnover of the enzyme. The structure of this transient protein complex has been studied using paramagnetic NMR spectroscopy. Gadolinium complexes were attached specifically through two engineered Cys residues on three sites on the surface of nitrite reductase, causing strong distance-dependent relaxation effects on the residues of pseudoazurin. Docking of the two proteins based on these NMR-derived distance restraints and the chemical shift perturbation data shows convergence to a cluster of structures with an average root-mean-square deviation of 1.5 Å. The binding interface consists of polar and non-polar residues surrounded by charges. The interprotein distance between the two type-1 copper sites is 15.5(± 0.5) Å, enabling fast interprotein electron transfer. The NMR-based lower limit estimate of 600 s−1 for the dissociation rate constant and the fast electron transfer are consistent with the transient nature of the complex. 相似文献
98.
This paper presents Computational fluid dynamic (CFD) analysis of blood flow in three different 3-D models of left coronary artery (LCA). A comparative study of flow parameters (pressure distribution, velocity distribution and wall shear stress) in each of the models is done for a non-Newtonian (Carreau) as well as the Newtonian nature of blood viscosity over a complete cardiac cycle. The difference between these two types of behavior of blood is studied for both transient and steady states of flow. Additionally, flow parameters are compared for steady and transient boundary conditions considering blood as non-Newtonian fluid. The study shows that the highest wall shear stress (WSS), velocity and pressure are found in artery having stenosis in all the three branches of LCA. The use of Newtonian blood model is a good approximation for steady as well as transient blood flow boundary conditions if shear rate is above 100 s-1. However, the assumption of steady blood flow results in underestimating the values of flow parameters such as wall shear stress, pressure and velocity. 相似文献
99.
Miklós Z Ivanics T Roemen TH van der Vusse GJ Dézsi L Szekeres M Kemecsei P Tóth A Kollai M Ligeti L 《Molecular and cellular biochemistry》2003,250(1-2):115-124
The main aim of this study was to assess the kinetics of intracellular free calcium (Ca2+
i) handling by isolated rat hearts rendered ischemic for 30 min followed by 30 min of reperfusion analyzing the upstroke and downslope of the Ca2+
i transient. Changes in mechanical performance and degradation of membrane phospholipids – estimated by tissue arachidonic acid content – were correlated with Ca2+
i levels of the heart. The fluorescence ratio technique was applied to estimate Ca2+
i. The disappearance of mechanical activity of the heart preceded that of the Ca2+
i transient in the first 2 min of ischemia. The slope of upstroke of the Ca2+
i transient, reflecting Ca2+ release, decreased by 60%, while the duration of the downslope of the transient, reflecting Ca2+ sequestration, expressed a significant prolongation (105 ± 17 vs. 149 ± 39 msec) during the first 3 min of ischemia. At about 20 min of ischemia end-diastolic pressure expressed a 3.5-fold increase (contracture) when the fluorescence ratio showed a 2-fold elevation. Reperfusion was accompanied with a further precipitous increase in end-diastolic pressure, while resting Ca2+
i remained at end-ischemic levels. Increases in the arachidonic acid (AA) content of the ischemic and postischemic hearts were proportional to Ca2+
i levels. In summary, the present findings indicate that both calcium release and removal are hampered during the early phase of ischemia. Moreover, a critical level of Ca2+
i and a critical duration of ischemia may exist to provoke contracture of the heart. Upon reperfusion the hearts show membrane phospholipid degradation and signs of stunning exemplified by elevated AA levels, partial recovery of Ca2+
i handling and sustained depression of mechanical performance. 相似文献
100.
Abstract Polydnaviruses are a group of insect DNA viruses and are characterized in their segmented genome that is located in the chromosome(s) of host wasps. A polydnavirus, Cotesia plutellae bracovirus (CpBV), encodes a viral ribonuclease (RNase) T2 in a specific segment #3 (CpBV‐S3). This study tested its effect on gene expression associated with host immune responses in the diamondback moth, Plutella xylostella. Micro‐injection of CpBV‐S3 into nonparasitized larvae induced expression of its two encoded genes, CpBV‐ORF301 (=CpBV‐RNase T2) and CpBV‐ORF302. In response to a bacterial challenge, four antimicrobial peptide genes (hemolin, gloverin, cecropin and lysozyme) and six phenoloxidase (PO)–associated genes (proPO‐activating proteinase, PO, serine proteinase homolog and serpins 1–3) were up‐regulated in their expressions. However, the transient expression of CpBV‐S3 suppressed the expressions of cecropin, PO and serpin 1. Double‐stranded RNA specific to the viral RNase T2 could specifically knockdown the viral gene expression and restored the three gene expressions suppressed in the larvae injected with CpBV‐S3. The inhibitory activity of the viral RNase T2 on the target genes was further proven by the suppression of PO activation in response to bacterial challenge in the larvae injected with CpBV‐S3. This immunosuppression by the expression of the viral RNase T2 resulted in significant increase of pathogen susceptibility of P. xylostella against Bacillus thuringiensis or baculovirus infection. 相似文献