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991.
Raffaele Lombardi Maria Elena Villani Mariasole Di Carli Patrizia Brunetti Eugenio Benvenuto Marcello Donini 《Transgenic research》2010,19(6):1083-1097
It was previously demonstrated that the tumour-targeting antibody mAb H10 can be transiently expressed and purified at high
levels in Nicotiana benthamiana by using a vacuum-agroinfiltration system boosted by the use of a virus silencing suppressor protein. Scope of this work
was to analyse different steps of protein extraction from agroinfiltrated leaves to optimise the purification process of the
secretory mAb H10 providing new insights in the field of large-scale plant production. Two different extraction procedures
(mechanical shearing/homogenisation and recovery of intercellular fluids -IFs-) were evaluated and compared in terms of purified
antibody yields, antibody degradation and total phenolic compounds content. Mechanical grinding from fresh leaf tissues gave
the highest purification yield (75 mg/kg Fresh Weight -75% intact tetrameric IgG-) and total phenolics concentration in the
range of 420 μg/g FW. The second extraction procedure, based on the recovery of IFs, gave purification yields of 15–20 mg/kg
FW (corresponding to 27% of total soluble protein) in which about 40% of purified protein is constituted by fully assembled
IgG with a total phenolic compounds content reduced by one order of magnitude (21 μg/g FW). Despite a higher antibody degradation,
purification from intercellular fluids demonstrated to be very promising since extraction procedures resulted extremely fast
and amenable to scaling-up. Overall data highlight that different extraction procedures can dramatically affect the proteolytic
degradation and quality of antibody purified from agroinfiltrated N. benthamiana leaves. Based on these results, we optimised a pilot-scale purification protocol using a two-step purification procedure
from batches of fresh agroinfiltrated leaves (250 g) allowing purification of milligram quantities (average yield 40 mg/kg
FW) of fully assembled and functional IgG with a 99.4% purity, free of phenolic and alkaloid compounds with low endotoxin
levels (<1 EU/ml). 相似文献
992.
Ju-Hong Zhang Li-Wen Sun Lin-Lin Liu Jie Lian Shao-Li An Xu Wang Jing Zhang Jun-Ling Jin Shan-Yu Li Jing-Hui Xi 《Plant Molecular Biology Reporter》2010,28(2):324-333
In this study, comparative proteomics was used to investigate the interaction of Spodoptera exigua and Arabidopsis thaliana. By using 2-D electrophoresis of differentially expressed proteins, combined with high-throughput matrix-assisted laser desorption/ionization
time of flight mass spectrometry (MALDI-TOF MS) and MALDI-TOF/TOF MS, the changes in the abundance of proteins induced by
insect feeding were studied in A. thaliana. More than 1,100 protein spots were reproducibly detected on each gel. The intensities of 30 protein spots in particular
changed significantly, showing differences in volume of at least twofold. Among these, 17 protein spots were upregulated,
and 13 were downregulated following an 8-h insect feeding period. Nineteen insect-feeding-responsive proteins were identified,
all of which were involved in metabolic regulation, binding functions or cofactor requirement of protein, cell rescue, and
defense and virulence, as assessed by Munich Information Center for Protein Sequences function category. About 50% of these
were involved in metabolism, including transketolase, S-adenosylmethionine synthase 3, 2,3-biphosphoglycerate-independent phosphoglycerate mutase, beta-ureidopropionase, GDP-d-mannose 3′,5′-epimerase, and fatty acid synthase. The identification of insect-feeding-responsive proteins on Arabidopsis provides not only new insights into insect stress but also a good start for further investigation of their functions. Understanding
how the plant responses to insects in the proteomic level will provide tools for a better management of insect pest in the
field. 相似文献
993.
Wei Fan Stefan A. W. Bouwense Ross Crawford Yin Xiao 《Journal of molecular histology》2010,41(1):51-60
Despite the important physiological role of periosteum in the pathogenesis and treatment of osteoporosis, little is known
about the structural and cellular characteristics of periosteum in osteoporosis. To study the structural and cellular differences
in both diaphyseal and metaphyseal periosteum of osteoporotic rats, samples from the right femur of osteoporotic and normal
female Lewis rats were collected and tissue sections were stained with hematoxylin and eosin, antibodies or staining kit against
tartrate resistant acid phosphatase (TRAP), alkaline phosphatase (ALP), vascular endothelial growth factor (VEGF), von Willebrand
(vWF), tyrosine hydroxylase (TH) and calcitonin gene-related peptide (CGRP). The results showed that the osteoporotic rats
had much thicker and more cellular cambial layer of metaphyseal periosteum compared with other periosteal areas and normal
rats (P < 0.001). The number of TRAP+ osteoclasts in bone resorption pits, VEGF+ cells and the degree of vascularization were found to be greater in the cambial layer of metaphyseal periosteum of osteoporotic
rats (P < 0.05), while no significant difference was detected in the number of ALP+ cells between the two groups. Sympathetic nerve fibers identified by TH staining were predominantly located in the cambial
layer of metaphyseal periosteum of osteoporotic rats. No obvious difference in the expression of CGRP between the two groups
was found. In conclusion, periosteum may play an important role in the cortical bone resorption in osteoporotic rats and this
pathological process may be regulated by the sympathetic nervous system. 相似文献
994.
Annexins belong to a multigene family of Ca2+ dependent, phospholipid and cytoskeleton binding proteins. They have been shown to be upregulated under various stress conditions.
We generated transgenic cotton plants expressing mustard annexin (AnnBj1), which showed enhanced tolerance towards different abiotic stress treatments like sodium chloride, mannitol, polyethylene
glycol and hydrogen peroxide. The tolerance to these treatments was associated with decreased hydrogen peroxide levels and
enhanced total peroxidase activity, enhanced content of osmoprotectants- proline and sucrose in transgenic plants. They showed
higher retention of total chlorophyll and reduced TBARS in leaf disc assays with stress treatments, and decreased hydrogen
peroxide accumulation in the stomatal guard cells when compared to their wild type counterparts. They also showed significantly
enhanced fresh weight, relative water content, dry weight under stress. Treatment with sodium chloride resulted in enhanced
expression of genes for ∆-pyrroline-5-carboxylase synthetase in leaves, and sucrose phosphate synthase, sucrose synthase and
cellulose synthase A in the leaves and fibers of transgenic plants. The transgenic plants maintained normal seed development,
fiber quality and cellulose content under stress. 相似文献
995.
Nazim Fatès 《Swarm Intelligence》2010,4(2):91-115
The decentralised gathering problem consists in grouping in a compact cluster agents that are initially randomly scattered.
We propose a bio-inspired algorithm, the Reaction–Diffusion–Chemotaxis aggregation scheme, to group agents that have limited
abilities. The agents and their environment are described with a stochastic model inspired by the aggregation of the Dictyostelium discoideum cellular slime mold. The environment is an active lattice, whose cells transmit information according to a reaction–diffusion
mechanism. The agents are virtual amoebae; they trigger excitations randomly and move by following reaction–diffusion waves.
We demonstrate that despite its simplicity, this model exhibits interesting properties of self-organisation and is efficient
for gathering agents. Moreover, observations show that the system is robust to various perturbations, such as the presence
of obstacles on the lattice or noise in the movements of the agents. 相似文献
996.
Silvano Fares Elina Oksanen Mika Lännenpää Riitta Julkunen-Tiitto Francesco Loreto 《Photosynthesis research》2010,104(1):61-74
Plants are exposed to increasing levels of tropospheric ozone concentrations. This pollutant penetrates in leaves through
stomata and quickly reacts inside leaves, thus making plants valuable ozone sinks, but at the same time triggers oxidation
processes which lead to leaf injuries. To counteract these negative effects, plants produce an array of antioxidants which
react with ozone and reactive molecules which ozone generates in the leaf tissues. In this study, we measured the effect of
an ozone concentration which is likely to be attained in many areas of the world in the near future (80 ppb) on leaves of
the vertical profile of the widespread agroforestry species Populus nigra. Changes in (1) physiological parameters (photosynthesis and stomatal conductance), (2) ozone uptake, (3) emission of volatile
organic compounds (VOCs, i.e. isoprene, methanol and other oxygenated compounds), (4) concentration of antioxidant surface
compounds, and (5) concentration of phenolic compounds were assessed. The aim was to assess whether the defensive pathways
leading to isoprenoids and phenolics formation were induced when a moderate and chronic increment of ozone is not able to
damage photosynthesis. No visual injuries and minor changes in physiology and ozone uptake were observed. The emission of
isoprene and oxygenated six-carbon (C6) volatiles were inhibited by ozone, whereas methanol emission was increased, especially
in developing leaves. We interpret these results as suggesting an ontogenetic shift in ozone-treated leaves, leading to a
slower development and a faster senescence. Most surface and phenolic compounds showed a declining trend in concentration
from the youngest to the fully expanded leaves. Ozone reduced the concentrations of chlorogenic acid derivatives at the leaf
surface, whereas in total leaf extracts a metabolic shift towards few phenolics with higher antioxidant capacity was observed. 相似文献
997.
Background
Both bovine coronavirus (BCV) and bovine respiratory syncytial virus (BRSV) infections are currently wide-spread in the Swedish dairy cattle population. Surveys of antibody levels in bulk tank milk have shown very high nationwide prevalences of both BCV and BRSV, with large variations between regions. In the Swedish beef cattle population however, no investigations have yet been performed regarding the prevalence and geographical distribution of BCV and BRSV. A cross-sectional serological survey for BCV and BRSV was carried out in Swedish beef cattle to explore any geographical patterns of these infections. 相似文献998.
The mammary gland undergoes hormonally controlled cycles of pubertal maturation, pregnancy, lactation, and involution, and
these processes rely on complex signaling mechanisms, many of which are controlled by cell–cell and cell–matrix adhesion.
The adhesion of epithelial cells to the extracellular matrix initiates signaling mechanisms that have an impact on cell proliferation,
survival, and differentiation throughout lactation. The control of integrin expression on the mammary epithelial cells, the
composition of the extracellular matrix and the presence of secreted matricellular proteins all contribute to essential adhesion
signaling during lactogenesis. In vitro and in vivo studies, including the results from genetically engineered mice, have
shed light on the regulation of these processes at the cell and tissue level and have led to increased understanding of the
essential signaling components that are regulated in temporal and cell specific manner during lactogenesis. Recent studies
suggest that a secreted matricellular protein, CTGF/CCN2, may play a role in lactogenic differentiation through binding to
β1 integrin complexes, enhancing the production of extracellular matrix components and contributions to cell adhesion signaling. 相似文献
999.
Nakazawa Y Sagane Y Kikuchi T Uchino M Nagai T Sato H Toeda K Takano K 《The protein journal》2010,29(8):598-608
We previously isolated Streptomyces racemochromogenes strain 10-3, which produces a phospholipase D (PLD) with high transphosphatidylation activity. Here, we purified and cloned
the PLD (PLD103) from the strain. PLD103 exerted the highest hydrolytic activity at a slightly alkaline pH, which is in contrast
to the majority of known Streptomyces PLDs that have a slightly acidic optimum pH. PLD103 shares only 71–76% amino acid sequence identity with other Streptomyces PLDs that have a slightly acidic optimum pH; thus, the diversity in the primary structure might explain the discrepancy observed
in the optimum pH. The purified PLD displayed high transphosphatidylation activity in the presence of glycerol, l-serine, and 2-aminoethanol hydrochloride with a conversion rate of 82–97% in a simple one-phase system, which was comparable
to the rate of other Streptomyces PLDs in a complicated biphasic system. 相似文献
1000.
Geert Potters Marcel A. K. Jansen Nele Horemans Yves Guisez Taras Pasternak 《In vitro cellular & developmental biology. Plant》2010,46(3):289-297
A Nicotiana tabacum L. SR-1 leaf protoplast system was used to study the effects of dehydroascorbate and glutathione on cellular development. We found that dehydroascorbate is readily taken up by protoplasts and internally reduced to ascorbate. Concomitantly, cell division was inhibited and cell expansion stimulated. In this respect, dehydroascorbate counteracted auxin-mediated leaf protoplast development. In contrast to the effects of dehydroascorbate, glutathione-induced cell dedifferentiation, and this effect is similar to that mediated by high auxin concentrations. We conclude that dehydroascorbate and glutathione affect the auxin-mediated regulation of cellular development. Therefore, the biological role of these compounds extends beyond stress tolerance and defense. 相似文献