Crystal structure of an early protein-RNA assembly complex of the signal recognition particle

The signal recognition particle (SRP) is a universally conserved ribonucleoprotein complex that mediates the cotranslational targeting of secretory and membrane proteins to cellular membranes. A crucial early step in SRP assembly in archaea and eukarya is the binding of protein SRP19 to specific sites on SRP RNA. Here we report the 1.8 angstrom resolution crystal structure of human SRP19 in complex with its primary binding site on helix 6 of SRP RNA, which consists of a stem-loop structure closed by an unusual GGAG tetraloop. Protein-RNA interactions are mediated by the specific recognition of a widened major groove and the tetraloop without any direct protein-base contacts and include a complex network of highly ordered water molecules. A model of the assembly of the SRP core comprising SRP19, SRP54, and SRP RNA based on crystallographic and biochemical data is proposed.     

The crystal structure of the signal recognition particle in complex with its receptor

Cotranslational targeting of membrane and secretory proteins is mediated by the universally conserved signal recognition particle (SRP). Together with its receptor (SR), SRP mediates the guanine triphosphate (GTP)-dependent delivery of translating ribosomes bearing signal sequences to translocons on the target membrane. Here, we present the crystal structure of the SRP:SR complex at 3.9 angstrom resolution and biochemical data revealing that the activated SRP:SR guanine triphosphatase (GTPase) complex binds the distal end of the SRP hairpin RNA where GTP hydrolysis is stimulated. Combined with previous findings, these results suggest that the SRP:SR GTPase complex initially assembles at the tetraloop end of the SRP RNA and then relocalizes to the opposite end of the RNA. This rearrangement provides a mechanism for coupling GTP hydrolysis to the handover of cargo to the translocon.     

A small viral RNA is required for in vitro packaging of bacteriophage phi 29 DNA

A small RNA of Bacillus subtilis bacteriophage phi 29 is shown to have a novel and essential role in viral DNA packaging in vitro. This requirement for RNA in the encapsidation of viral DNA provides a new dimension of complexity to the attendant protein-DNA interactions. The RNA is a constituent of the viral precursor shell of the DNA-packaging machine but is not a component of the mature virion. Studies of the sequential interactions involving this RNA molecule are likely to provide new insight into the structural and possible catalytic roles of small RNA molecules. The phi 29 assembly in extracts and phi 29 DNA packaging in the defined in vitro system were strongly inhibited by treatment with the ribonucleases A or T1. However, phage assembly occurred normally in the presence of ribonuclease A that had been treated with a ribonuclease inhibitor. An RNA of approximately 120 nucleotides co-purified with the phi 29 precursor protein shell (prohead), and this particle was the target of ribonuclease action. Removal of RNA from the prohead by ribonuclease rendered it inactive for DNA packaging. By RNA-DNA hybridization analysis, the RNA was shown to originate from a viral DNA segment very near the left end of the genome, the end packaged first during in vitro assembly.     

Crystal structure of the ribonucleoprotein core of the signal recognition particle

The signal recognition particle (SRP), a protein-RNA complex conserved in all three kingdoms of life, recognizes and transports specific proteins to cellular membranes for insertion or secretion. We describe here the 1.8 angstrom crystal structure of the universal core of the SRP, revealing protein recognition of a distorted RNA minor groove. Nucleotide analog interference mapping demonstrates the biological importance of observed interactions, and genetic results show that this core is functional in vivo. The structure explains why the conserved residues in the protein and RNA are required for SRP assembly and defines a signal sequence recognition surface composed of both protein and RNA.     

Relationship between clinical signs and transmission of an infectious disease and the implications for control

Control of many infectious diseases relies on the detection of clinical cases and the isolation, removal, or treatment of cases and their contacts. The success of such "reactive" strategies is influenced by the fraction of transmission occurring before signs appear. We performed experimental studies of foot-and-mouth disease transmission in cattle and estimated this fraction at less than half the value expected from detecting virus in body fluids, the standard proxy measure of infectiousness. This is because the infectious period is shorter (mean 1.7 days) than currently realized, and animals are not infectious until, on average, 0.5 days after clinical signs appear. These results imply that controversial preemptive control measures may be unnecessary; instead, efforts should be directed at early detection of infection and rapid intervention.     

The structure of sister minichromosome DNA before anaphase in Saccharomyces cerevisiae

The role of DNA topology in holding sister chromatids together before anaphase was investigated by analyzing the structure of a small circular minichromosome in cell cycle (cdc) mutants of the yeast Saccharomyces cerevisiae. In the majority of cells arrested after S phase but before anaphase, sister minichromosome molecules are not topologically interlocked with each other. The analysis of the ploidy of minichromosomes in cells that are released from arrest demonstrates that the sister molecules are properly segregated when the cell cycle block is removed. Therefore, sister minichromosome molecules need not remain topologically interlocked until anaphase in order to be properly segregated, and topological interlocking of sister DNA molecules apparently is not the primary force holding sister chromatids together.     

The mechanism of DNA transfer in the mating system of an archaebacterium

The genetic transfer system in the extremely halophilic archaebacterium Halobacterium volcanii is the only archaebacterial mating system known. The mechanism of genetic transfer of this archaebacterium was studied by using the immobile plasmids pHV2 and pHV11 as cytoplasmic markers. It was found that the cytoplasms of the parental types do not mix during the mating process, that each parental type can serve both as a donor and as a recipient, and that cytoplasmic bridges, with dimensions of up to 2 micrometers long and 0.1 micrometer in diameter, were formed between the parental types. These bridges appear to be used for the transfer of DNA from one cell to another. If so, this archaebacterial mating system is different from both eubacterial conjugation and eukaryotic sexual cell fusion.     

Skinny hedgehog, an acyltransferase required for palmitoylation and activity of the hedgehog signal

One of the most dominant influences in the patterning of multicellular embryos is exerted by the Hedgehog (Hh) family of secreted signaling proteins. Here, we identify a segment polarity gene in Drosophila melanogaster, skinny hedgehog (ski), and show that its product is required in Hh-expressing cells for production of appropriate signaling activity in embryos and in the imaginal precursors of adult tissues. The ski gene encodes an apparent acyltransferase, and we provide genetic and biochemical evidence that Hh proteins from ski mutant cells retain carboxyl-terminal cholesterol modification but lack amino-terminal palmitate modification. Our results suggest that ski encodes an enzyme that acts within the secretory pathway to catalyze amino-terminal palmitoylation of Hh, and further demonstrate that this lipid modification is required for the embryonic and larval patterning activities of the Hh signal.     

FMDV P1基因DNA疫苗构建及免疫实验

[目的]构建口蹄疫DNA疫苗[方法]构建以PRV为载体的表达FMDV P1基因的质粒pIESZP1和pUTK3CP1,免疫小鼠并检测了免疫后小鼠的抗体水平.将构建的2个真核表达质粒分别转染Vero细胞,通过PCR、IFA、Western-blot等方法检测目的基因的转录和表达.将正确表达目的基因的DNA质粒肌肉接种3周龄的Balb/C小鼠,采用ELISA和SN方法检测了免疫小鼠体内的抗体水平.[结果]携带有FMDV衣壳蛋白P1基因的DNA质粒能引起小鼠产生特异性的体液免疫反应,两重组质粒诱导小鼠产生抗FMDV的抗体水平无差异.[结论]该研究为含有FMDV P1基因重组PRV以及重组病毒的动物免疫试验以评价基因工程疫苗的免疫效果奠定了基础.     

Reaction of the antitumor antibiotic CC-1065 with DNA: structure of a DNA adduct with DNA sequence specificity

Sequence-dependent variations in DNA revealed by x-ray crystallographic studies have suggested that certain DNA-reactive drugs may react preferentially with defined sequences in DNA. Drugs that wind around the helix and reside within one of the grooves of DNA have perhaps the greatest chance of recognizing sequence-dependent features of DNA. The antitumor antibiotic CC-1065 covalently binds through N-3 of adenine and resides within the minor groove of DNA. This drug overlaps with five base pairs for which a high sequence specificity exists.     

航空HF电台测试系统跳频信号源的实现

航空HF电台是航空通信系统的重要组成部分,担负着空空、空地之间的远程通信,关系到空空、空地之间的通信安全。通过分析航空HF电台测试和控制需求,提出基于"DDS+PLL"的检测仪跳频信号源的设计方案,介绍跳频信号源中频率合成单元电路工作原理和工作流程,分析跳频信号源各频率合成单元的输出频率的计算方法。测试结果表明,该跳频信号源具有频率稳定度高、频率分辨率高、抗干扰能力强和频率转换时间短等优点。     

The crystal structure of TAL effector PthXo1 bound to its DNA target

DNA recognition by TAL effectors is mediated by tandem repeats, each 33 to 35 residues in length, that specify nucleotides via unique repeat-variable diresidues (RVDs). The crystal structure of PthXo1 bound to its DNA target was determined by high-throughput computational structure prediction and validated by heavy-atom derivatization. Each repeat forms a left-handed, two-helix bundle that presents an RVD-containing loop to the DNA. The repeats self-associate to form a right-handed superhelix wrapped around the DNA major groove. The first RVD residue forms a stabilizing contact with the protein backbone, while the second makes a base-specific contact to the DNA sense strand. Two degenerate amino-terminal repeats also interact with the DNA. Containing several RVDs and noncanonical associations, the structure illustrates the basis of TAL effector-DNA recognition.     

电商环境下八马茶业的包装设计及其品牌营造

在电子商务营销成为茶叶销售重要渠道的当下,探究八马茶业是如何主动适应这种新环境,如何不断探索茶叶包装设计的改进及通过企业视觉识别设计和包装设计等进行品牌营造的。从电商环境下茶叶销售方式的改变,引出电商环境下茶业企业的标志设计和包装设计是消费者了解商品最重要的渠道。同时从八马茶业包装设计的材质、色彩、图案和文字等方面分析其包装设计的特点,并从企业命名、标志设计和包装设计等方面分析八马茶业打造企业品牌的途径。研究表明,八马茶业包装设计材料工艺讲究,运用中国传统的图案、色彩和文字,显得既具有历史厚重感,又兼具现代风格的简约性和可辨识性。八马茶业通过企业视觉识别系统和独特的包装设计,打造了独树一帜的"八马"品牌形象。     

The structure of rat preproatrial natriuretic factor as defined by a complementary DNA clone

The structure of rat preproatrial natriuretic factor ( preproANF ) was determined by nucleotide sequence analysis of an ANF complementary DNA clone. PreproANF is composed of a hydrophobic leader segment (20 amino acids), a precursor containing one glycosylation site (106 amino acids), and ANF (24 amino acids). Atrial natriuretic factor is located at the carboxyl terminus of the precursor molecule. The human, mouse, and rat genomes each contain a single ANF gene which is highly conserved.     

Chromatin structure and de novo methylation of sperm DNA: implications for activation of the paternal genome

The chromatin structure characteristic of constitutively expressed genes, tissue-specific genes, and inactive genes is absent in chicken sperm chromatin. However, point sites of undermethylation in sperm DNA within constitutively expressed genes, but not within globin genes or an inactive gene, correspond to the location of regions of altered chromatin structure (hypersensitive sites) in somatic tissue and spermatogonial cells. A de novo methylation process whereby regions within and flanking these genes become methylated, but which excludes the methylation of sequences within hypersensitive sites, occurs between the spermatogonial stage and the first meiotic prophase. These undermethylated regions may play a role in the activation of the paternal genome during embryogenesis.     

Crystal structure of the human two-pore domain potassium channel K2P1

Two-pore domain potassium (K(+)) channels (K2P channels) control the negative resting potential of eukaryotic cells and regulate cell excitability by conducting K(+) ions across the plasma membrane. Here, we present the 3.4 angstrom resolution crystal structure of a human K2P channel, K2P1 (TWIK-1). Unlike other K(+) channel structures, K2P1 is dimeric. An extracellular cap domain located above the selectivity filter forms an ion pathway in which K(+) ions flow through side portals. Openings within the transmembrane region expose the pore to the lipid bilayer and are filled with electron density attributable to alkyl chains. An interfacial helix appears structurally poised to affect gating. The structure lays a foundation to further investigate how K2P channels are regulated by diverse stimuli.     

用于RAPD分析的树莓基因组总DNA提取初报

以树莓嫩叶为材料,进行树莓基因组总DNA不同提取方法的比较试验.结果表明:CTAB法、SDS法、改良CTAB法均可成功提取树莓DNA,但无论在质量上还是在数量上,改良CTAB法都优于常规CTAB法和SDS法,所提取的DNA大多呈现无色透明状,A260/ A280值多数介于1.65～1.90,完全可以用于RAPD分析.