Cyclic AMP phosphodiesterase in cloned astrocytoma cells: norepinephrine induces a specific enzyme form

The soluble supernatant fraction of homogenates of cloned rat astrocytoma cells (linle C-2A) was subjected to polyacrylamide gel electrophoresis. Two peaks of adenosine 3',5'-monophosphate phosphodiesterase activity were found, corresponding to peaks I and IV of a similarly prepared homogenate of rat brain. Incubating cells with norepinephrine (0.3 mullimolar) caused about a threefold increase in the activity of peak IV but no change in peak I. This increase was completely inhibited by prior inclubation with propranolol (0.1 millimnolar), a beta-adrenergic blocking agent, or with cyclohexamnine (40 micromolar). a protein synthesis inhibitor. Induction of a specific phosphodiesterase form by norepinephrine sitggests another feedback control mechanism whereby an organism can prevent the effects of excessive sympathetic activity.     

Chemical conversion of a DNA-binding protein into a site-specific nuclease

The tryptophan gene (trp) repressor of Escherichia coli has been converted into a site-specific nuclease by covalently attaching it to the 1,10-phenanthroline-copper complex. In its cuprous form, the coordination complex with hydrogen peroxide as a coreactant cleaves DNA by oxidatively attacking the deoxyribose moiety. The chemistry for the attachment of 1,10-phenanthroline to the trp repressor involves modification of lysyl residues with iminothiolane followed by alkylation of the resulting sulfhydryl groups with 5-iodoacetamido-1,10-phenanthroline. The modified trp repressor cleaves the operators of aroH and trpEDCBA upon the addition of cupric ion and thiol in a reaction dependent on the corepressor L-tryptophan. Scission was restricted to the binding site for the repressor, defined by deoxyribonuclease I footprinting. Since DNA-binding proteins have recognition sequences approximately 20 base pairs long, the nucleolytic activities derived from them could be used to isolate long DNA fragments for sequencing or chromosomal mapping.     

Identification of p53 as a sequence-specific DNA-binding protein

The tumor-suppressor gene p53 is altered by missense mutation in numerous human malignancies. However, the biochemical properties of p53 and the effect of mutation on these properties are unclear. A human DNA sequence was identified that binds specifically to wild-type human p53 protein in vitro. As few as 33 base pairs were sufficient to confer specific binding. Certain guanines within this 33-base pair region were critical, as methylation of these guanines or their substitution with thymine-abrogated binding. Human p53 proteins containing either of two missense mutations commonly found in human tumors were unable to bind significantly to this sequence. These data suggest that a function of p53 may be mediated by its ability to bind to specific DNA sequences in the human genome, and that this activity is altered by mutations that occur in human tumors.     

Three-dimensional solution structure of a single zinc finger DNA-binding domain

The three-dimensional solution structure of a zinc finger nucleic acid binding motif has been determined by nuclear magnetic resonance (NMR) spectroscopy. Spectra of a synthetic peptide corresponding to a single zinc finger from the Xenopus protein Xfin yielded distance and dihedral angle constraints that were used to generate structures from distance geometry and restrained molecular dynamics calculations. The zinc finger is an independently folded domain with a compact globular structure in which the zinc atom is bound by two cysteine and two histidine ligands. The polypeptide backbone fold consists of a well-defined helix, starting as alpha and ending as 3(10) helix, packed against two beta strands that are arranged in a hairpin structure. A high density of basic and polar amino acid side chains on the exposed face of the helix are probably involved in DNA binding.     

水稻PHD锌指蛋白cDNA的克隆及其表达分析

利用cDNA-AFLP分析籼稻明恢86应答稻纵卷叶螟取食基因差异表达,发现1个与植物同源结构域(PHD)锌指蛋白高度同源的TDF,分离获得该TDF对应的粳稻全长cDNA.该全长cDNA与籼稻、玉米、蓖麻、葡萄、拟南芥和大豆等作物PHD锌指蛋白推定氨基酸序列的同源性分别为98.43%、86.01%、54.58%57.02...     

Encephalitogenic protein: structure

Amino acid sequences of encephalitogenic proteins from bovine cord and rabbit brain are reported. The bovine protein contains 45 residues. The rabbit protein is identical except for two isopolar substitutions, a dipeptide and amino acid deletion. Analysis of this protein and a 140-residue myelin basic protein indicates that the smaller protein is a portion of the larger encephalitogen. The larger myelin protein contains at least two encephalitogenic sites.     

Cyclic uridine-3',5'-phosphate: molecular structure

The crystal structure of the triethylammonium salt of cyclic uridine-3',5'-phosphate was solved by use of the tangent formula to refine phase angles based upon the positions of six of the atoms. The two independent uracil rings are planar and in the keto form. The base-sugar torsion angles are in the anti range. The sugar puckering is C3'-endo, and the ribose conformation about the C5'-C4' bond is transgauche.     

Metalloregulatory DNA-binding protein encoded by the merR gene: isolation and characterization

The MerR protein mediates the induction of the mercury resistance phenotype in bacteria; it has been isolated in order to study the effects of metal-ion induced changes in the metabolism of prokaryotic cells at the molecular level. After DNA sequences responsible for negative autoregulation were removed, the 16-kilodalton protein was overproduced and purified to more than 90 percent homogeneity by a salt extraction procedure that yields about 5 milligrams of protein per gram of cells. Complementation data, amino terminal analysis, gel filtration, and deoxyribonuclease I protection studies demonstrate that the purified merR gene product is a dimer under nondenaturing conditions and that it binds specifically to DNA, in the presence and absence of mercury, at a palindromic site which is directly between the -10 and -35 regions of the structural genes and adjacent to its own promoter. These initial results indicate that MerR is a DNA-binding metalloregulatory protein that plays a central role in this heavy metal responsive system and they delineate an operator site in the mer operon.     

Cyclic cytidine 2',3'-phosphate: molecular structure

Monoclinic crystals of the sodium salt of cytidine 2',3'-phosphate contain two anions in the asymmetric unit. Both bases are in the syn conformation, and the nucleotides are stacked together into an antiparallel stranded ribbon with the bases 3.3 angstroms apart. One ribose ring is planar, and the other has oxygen-1' puckered toward carbon-5'. The phosphorus atoms in the five-membered ester rings are puckered toward the sugars. The conformations about the carbon-4'-carbon-5' bonds are gauche-trans and gauche-gauche.     

Aromatic-aromatic interaction: a mechanism of protein structure stabilization

Analysis of neighboring aromatic groups in four biphenyl peptides or peptide analogs and 34 proteins reveals a specific aromatic-aromatic interaction. Aromatic pairs (less than 7 A between phenyl ring centroids) were analyzed for the frequency of pair type, their interaction geometry (separation and dihedral angle), their nonbonded interaction energy, the secondary structural locations of interacting residues, their environment, and their conservation in related molecules. The results indicate that on average about 60 percent of aromatic side chains in proteins are involved in aromatic pairs, 80 percent of which form networks of three or more interacting aromatic side chains. Phenyl ring centroids are separated by a preferential distance of between 4.5 and 7 A, and dihedral angles approaching 90 degrees are most common. Nonbonded potential energy calculations indicate that a typical aromatic-aromatic interaction has energy of between -1 and -2 kilocalories per mole. The free energy contribution of the interaction depends on the environment of the aromatic pair. Buried or partially buried pairs constitute 80 percent of the surveyed sample and contribute a free energy of between -0.6 and -1.3 kilocalories per mole to the stability of the protein's structure at physiologic temperature. Of the proteins surveyed, 80 percent of these energetically favorable interactions stabilize tertiary structure, and 20 percent stabilize quaternary structure. Conservation of the interaction in related molecules is particularly striking.     

一种基于树的蛋白质功能预测算法：KDE–CSSA

针对在每个标签类上直接学习分类模型计算代价高和树层次中低层结点训练数据扭曲的问题,提出了一种基于树层次的蛋白质功能预测算法:核依赖估计–压缩排序选择算法(KDE–CSSA)。该算法先将标签向量投影到标签核的主成分上,仅仅学习少量的回归模型,然后将预测的数值向量投影回原来标签向量空间,利用压缩排序和选择算法获取满足树属性的0,1标签向量。在12个基因组数据集上使用精确率和召回率作为评测标准的实验结果表明,KDE–CSSA算法性能优于目前优秀的CLUS–HMC算法。     

Isolation of an olfactory cDNA: similarity to retinol-binding protein suggests a role in olfaction

Molecular cloning techniques were used to isolate and characterize a protein possibly involved in the signal transducing system in olfactory tissue of the frog Rana pipiens. A complementary DNA library was constructed with messenger RNA obtained from frog olfactory neuroepithelium. A 700-base pair complementary DNA clone encoding a protein with a molecular weight of 20,300 was identified by differential hybridization analysis with polyadenylated RNA from olfactory epithelium and nonsensory respiratory epithelium. The messenger RNA corresponding to this clone was abundant in the cells of Bowman's glands in olfactory tissue but not in respiratory epithelium nor in several other tissues. The predicted sequence of this protein is homologous to members of a family of proteins that bind and transport small molecules in serum, suggesting that this protein may also bind and transport odorants in the mucus secreted by Bowman's glands.     

Brain glutamate decarboxylase cloned in lambda gt-11: fusion protein produces gamma-aminobutyric acid

Glutamate decarboxylase (GAD; E.C. 4.1.1.15) converts glutamate to gamma-aminobutyric acid (GABA), the major inhibitory neurotransmitter in the vertebrate central nervous system. This report describes the isolation of a GAD complementary DNA clone by immunological screening of a lambda gt-11 brain complementary DNA expression library. The fusion protein produced by this clone catalyzes the conversion of glutamate to GABA and carbon dioxide, confirming its identity as GAD. Antibodies to beta-galactosidase remove GAD enzymatic activity from solution, showing that this activity is associated with the fusion protein. In immunoblotting experiments all three available antisera to GAD reacted with the fusion polypeptide and with two major polypeptides (molecular size, 60,000 and 66,000 daltons) in brain extracts.     

Immunoglobulin structure: partial amino acid sequence of a Bence Jones protein

Sequence analysis and ordering of the soluble tryptic peptides of one Bence Jones protein and comparison with partial sequence data for another have revealed many structural differences in the half of the molecule with the terminal amino group, but only one structural difference in the half of the molecule having the terminal carboxyl group. Somatic chromosomal rearrangements may effect such changes and account for variability in antibody structure.     

牦牛PLIN2基因的克隆及其在卵巢不同发情阶段的表达

\t\t\t\t\t目的\t\t\t\t\t揭示PLIN2基因对宣和猪生长性状的影响。\t\t\t\t\t\t\t\t\t\t\t\t\t\t\t\t\t\t方法\t\t\t\t\t以357头宣和猪为试验材料,采用PCR产物直接测序法检测了PLIN2基因的5′-UTR和3′-UTR多态性,分析了各SNP位点不同基因型的生长性状差异。\t\t\t\t\t\t\t\t\t\t\t\t\t\t\t\t\t\t结果\t\t\t\t\t在5′-UTR检测到2个SNP位点(G158A、T491G)、3′-UTR检出1个SNP位点(G5689A),上述3个SNP位点分别以GG、TT和GG基因型频率最高,G、T和G为优势等位基因;除G158A位点未达到Hardy-Weinberg平衡状态(P<0.01)外,其余2个突变位点均处于平衡状态(P>0.05),且杂合度较高、遗传多样性较为丰富。在6月龄体重、4—6月龄日增重和70日龄—6月龄日增重上,G158A位点的GG型、T491G位点的TT型和G5689A位点的GG型均显著高于同位点其他基因型(P<0.01或P<0.05);且3个SNP位点不同单倍型组合对生长性状的影响呈现了明显的协同作用,即GTG/GTG组合的6月龄体重、4—6月龄日增重和70日龄—6月龄日增重最高(P<0.01或P<0.05)。\t\t\t\t\t\t\t\t\t\t\t\t\t\t\t\t\t\t结论\t\t\t\t\t研究结果从PLIN2基因多态性角度印证了宣和猪新品种选育工作的有效性,初步证实PLIN2基因与宣和猪生长性状间存在显著关联。\t\t\t\t\t\t\t\t\t     

Creation of a protein vector construct including an SSBTne DNA-binding domain and VirD2 nuclear localization signal

A genetic engineering construct is created which encodes a chimeric protein for nonviral transgenesis that includes the SSB DNA-binding domain and VirD2 nuclear localization signal. A method is developed for purifying the target protein.