菜心中SDS-PAGE向负极泳动的富含苯丙氨酸的蛋白的发现

在提取乙醇酸氧化酶同工酶的过程中, 发现了一种在SDS变性下向负极泳动的富含苯丙氨酸的碱性蛋白.在3次重复实验中, 均可以从SDS-PAGE后的负极缓冲液中获得这种蛋白, 其碱性与酸性氨基酸残基的比例分别高达1.46、1.28和0.89,而苯丙氨酸含量分别高达为60.00%、65.62%和62.30%.其余氨基酸残基的含量则和普通蛋白的相近似.由于该富含苯丙氨酸的碱性蛋白是水溶性的, 碱性氨基酸残基应主要分布在蛋白的表面,而苯丙氨酸则组成一个强疏水内核.     

Stage-dependent Dishevelled-1 expression during mouse spermatogenesis suggests a role in regulating spermatid morphological changes

Dishevelled (Dsh in Drosophila or DVL in mice) is a member of the highly conserved Wg/Wnt signaling pathway, which regulates important processes such as cell proliferation, polarity, and specification of cell fate. Three orthologous genes of Dishevelled (Dvl-1, Dvl-2, and Dvl-3) have been found in both humans and mice. They play pivotal roles in regulating cell morphology and a variety of changes in cell behaviors. In the present study, we show that the expression of Dvl-1 is stage-dependent during mouse spermatogenesis, although Dvl-2 and Dvl-3 show relative consistent expression. The expression of Dvl-1 mRNA first appears in pachytene spermatocytes, increases in round and elongating spermatids, and then turns to an undetectable level in mature sperm cells. Analyses of immunohistochemistry and immunofluorescence staining show that DVL-1 is present diffusely in the cytoplasm of pachytene spermatocytes and exhibits mainly a vesicular pattern and perinuclear distribution and a weak diffusely cytoplasmic signal in round and elongating spermatids. The vesicular pattern of DVL-1 has been observed by previous studies in somatic cells, and suggested to play roles in signal transduction. Immunoprecipitation experiments show that DVL-1 coimmunprecipitates with spermatogenic cells beta-actin rather than alpha-tubulin. These results indicate that DVL-1 may be involved in spermatid morphological changes during mouse spermiogenesis through mediating signal transduction and/or regulating actin cytoskeleton organization.     

Capsid-like arrays in crystals of chimpanzee adenovirus hexon

The major coat protein, hexon, from a chimpanzee adenovirus (AdC68) is of interest as a target for vaccine vector modification. AdC68 hexon has been crystallized in the orthorhombic space group C222 with unit cell dimensions of a = 90.8 A, b = 433.0 A, c = 159.3 A, and one trimer (3 x 104,942 Da) in the asymmetric unit. The crystals diffract to 2.1 A resolution. Initial studies reveal that the molecular arrangement is quite unlike that in hexon crystals for human adenovirus. In the AdC68 crystals, hexon trimers are parallel and pack closely in two-dimensional continuous arrays similar to those formed on electron microscope grids. The AdC68 crystals are the first in which adenovirus hexon has molecular interactions that mimic those used in constructing the viral capsid.     

Proteome analysis of human liver carcinoma Huh7 cells harboring hepatitis C virus subgenomic replicon

Chronic infection by hepatitis C virus (HCV) is closely correlated with serious liver diseases. Although considerable progress has been made during recent years, the mechanism of replication and pathogenesis of HCV infection are still elusive. We have applied proteomic techniques in this work to globally analyze the protein expression profiles of a human liver cell lines Huh7 in absence and presence of HCV replication, aiming at elucidating the components of HCV replication and the cellular responses to HCV replication. The protein mixtures of three subcellular fractions from Huh7 and Huh7-HCV were separated by 2-DE under various pH gradients. Differentially expressed spots were identified by MALDI-TOF MS, followed by database searching. A total of 179 comparative proteins were identified unambiguously, including proteins associated with host cytoskeleton, intracellular traffic, oxidative and ER stress, proteasome degradation, translation, apoptosis, proliferation, etc. Host proteins known to interact with HCV proteins, such as HSP27, alpha-actinin, nucleolin and eukaryotic initiation factor 4A-I, were elevated in Huh7-HCV cells. Our study provides the global information of proteomic alteration of Huh7 cells in the presence of HCV replication and the clues for further understanding of the mechanism of HCV replication and pathogenesis.     

Diversity of centromeric repeats in two closely related wild rice species, Oryza officinalis and Oryza rhizomatis

Oryza officinalis (CC, 2n=24) and Oryza rhizomatis (CC, 2n=24) belong to the Oryza genus, which contains more than 20 identified wild rice species. Although much has been known about the molecular composition and organization of centromeres in Oryza sativa, relatively little is known of its wild relatives. In the present study, we isolated and characterized a 126-bp centromeric satellite (CentO-C) from three bacterial artificial chromosomes of O. officinalis. In addition to CentO-C, low abundance of CentO satellites is also present in O. officinalis. In order to determine the chromosomal locations and distributions of CentO-C (126-bp), CentO (155 bp) and TrsC (366 bp) satellite within O. officinalis, fluorescence in situ hybridization examination was done on pachytene or metaphase I chromosomes. We found that only ten centromeres (excluding centromere 7 and 2) contain CentO-C arrays in O. officinalis, while centromere 7 comprises CentO satellites, and centromere 2 is devoid of any detectable satellites. For TrsC satellites, it was detected at multiple subtelomeric regions in O. officinalis, however, in O. rhizomatis, TrsC sequences were detected both in the four centromeric regions (CEN 3, 4, 10, 11) and the multiple subtelomeric regions. Therefore, these data reveal the evolutionary diversification pattern of centromere DNA within/or between close related species, and could provide an insight into the dynamic evolutionary processes of rice centromere.     

Effects of the infestation by Actinote thalia pyrrha (Fabricius) on the physiological indexes of Mikania micrantha leaves

Mikania micrantha is a dangerous exotic weed now found throughout Guangdong Province, China. Introduced by the Indonesian Oil Palm Institute, lepidopteran defoliator Actinote thalia pyrrha (Fabricius) is a potential biological control factor for M. micrantha. Changes in the activities of superoxide dismutase (SOD), catalase (CAT), peroxidase (POD), polyphenoldoxidase (PPO), the total phenolics content, and the capacity of scavenging 1,1-diphenyl-2-picrylhydazyl (DPPH) free radicals in M. micrantha leaves were analyzed after infestation by A. thalia pyrrha larvae. The results showed that a slight change of all the indexes in the damaged leaves appeared within 3 h. After four days of infestation by A. thalia pyrrha , SOD and POD activities in the damaged leaves were higher than those of the control. But CAT activity was lower than that of the control, indicating that the SOD and POD functions were impeded more than those of CAT. The SOD and POD activities reached their highest values after 48 h of infestation by A. thalia pyrrha and then decreased, whereas the highest activity of CAT occurred after 24 h and then decreased. PPO activity fluctuated greatly with its three peak values being 1.83, 1.92, and 2.17 times that of the control. The total phenolics content increased initially and then decreased after 72 h of infestation by A. thalia pyrrha . The capacity of scavenging DPPH was significantly lower than that of the control (p<0.05). SOD activity positively correlated with CAT and POD activities for both damaged and normal leaves. However, the value of the correlation coefficient in the damaged leaves was higher than that in the normal leaves. The total phenolics content also showed a weak positive correlation with PPO activity. It is suggested that the protective response of M. micrantha to infestation by A. thalia pyrrha was short and limited, but resulted in a reduced function of the leaves, disturbed the metabolism in the protective enzyme system, and decreased the antioxidative capacity. M. micrantha was unable to resist the feeding stress by altering the content of the total phenolics.