Crystal structure of 15-mer DNA duplex containing unpaired bases

Errors during DNA replication or repair can lead to the presence of unpaired or inserted bases in the double helix, as well as to mismatched base pairs. So far only structures of the latter type have been characterized by X-ray crystallography. We report here a 3-A crystal structure of DNA 15-mer d(CGCGAAATTTACGCG), which forms a duplex with two unpaired adenine residues looped outside the B-type helix. This arrangement is in disagreement with the nuclear magnetic resonance spectroscopy results for the same 15-mer in solution, indicating polymorphic nature of the structure adopted by this sequence.     

Structure of an adenine-cytosine base pair in DNA and its implications for mismatch repair

Mutational pathways rely on introducing changes in the DNA double helix. This may be achieved by the incorporation of a noncomplementary base on replication or during genetic recombination, leading to substitution mutation. In vivo studies have shown that most combinations of base-pair mismatches can be accommodated in the DNA double helix, albeit with varying efficiencies. Fidelity of replication requires the recognition and excision of mismatched bases by proofreading enzymes and post-replicative mismatch repair systems. Rates of excision vary with the type of mismatch and there is some evidence that these are influenced by the nature of the neighbouring sequences. However, there is little experimental information about the molecular structure of mismatches and their effect on the DNA double helix. We have recently determined the crystal structures of several DNA fragments with guanine X thymine and adenine X guanine mismatches in a full turn of a B-DNA helix and now report the nature of the base pairing between adenine and cytosine in an isomorphous fragment. The base pair found in the present study is novel and we believe has not previously been demonstrated. Our results suggest that the enzymatic recognition of mismatches is likely to occur at the level of the base pairs and that the efficiency of repair can be correlated with structural features.     

Molecular structure of r(GCG)d(TATACGC): a DNA--RNA hybrid helix joined to double helical DNA

The molecule r(GCG)d(TATACGC) is self-complementary and forms two DNA--RNA hybrid segments surrounding a central region of double helical DNA; its molecular structure has been solved by X-ray analysis. All three parts of the molecule adopt a conformation which is close to that seen in the 11-fold RNA double helix. The conformation of the ribonucleotides is partly determined by water molecules bridging between the ribose O2' hydroxyl group and cytosine O2. The hybrid-DNA duplex junction contains no structural discontinuities. However, the central DNA TATA sequence has some structural irregularities.     

Recognition of DNA damage by the Rad4 nucleotide excision repair protein

Mutations in the nucleotide excision repair (NER) pathway can cause the xeroderma pigmentosum skin cancer predisposition syndrome. NER lesions are limited to one DNA strand, but otherwise they are chemically and structurally diverse, being caused by a wide variety of genotoxic chemicals and ultraviolet radiation. The xeroderma pigmentosum C (XPC) protein has a central role in initiating global-genome NER by recognizing the lesion and recruiting downstream factors. Here we present the crystal structure of the yeast XPC orthologue Rad4 bound to DNA containing a cyclobutane pyrimidine dimer (CPD) lesion. The structure shows that Rad4 inserts a beta-hairpin through the DNA duplex, causing the two damaged base pairs to flip out of the double helix. The expelled nucleotides of the undamaged strand are recognized by Rad4, whereas the two CPD-linked nucleotides become disordered. These findings indicate that the lesions recognized by Rad4/XPC thermodynamically destabilize the Watson-Crick double helix in a manner that facilitates the flipping-out of two base pairs.     

Structure refined to 2A of a nicked DNA octanucleotide complex with DNase I

The cutting rates of bovine pancreatic deoxyribonuclease I (DNase I) vary along a given DNA sequence, indicating that the enzyme recognizes sequence-dependent structural variations of the DNA double-helix. In an attempt to define the helical parameters determining this sequence-dependence, we have co-crystallized a complex of DNase I with a self-complementary octanucleotide and refined the crystal structure at 2 A resolution. This structure confirms the basic features of an early model, namely that an exposed loop of DNase I binds in the minor groove of B-type DNA and that interactions do occur with the backbone of both strands. Nicked octamer duplexes that have lost a dinucleotide from the 3'-end of one strand are hydrogen-bonded across a two-fold axis in the crystal to form a quasi-continuous double helix of 14 base pairs. The DNA 14-mer has a B-type conformation and shows substantial distortion of both local and overall helix parameters, induced mainly by the tight interaction of Y73 and R38 in the unusually wide minor groove. Directly coupled to the widening of the groove by approximately 3A is a 21.5 degree bend of the DNA away from the bound enzyme towards the major groove, suggesting that both DNA stiffness and groove width are important in determining the sequence-dependence of the enzyme cutting rate. A second cut of the DNA which is induced by diffusion of Mn2+ into the co-crystals suggests that there are two active sites in DNase I separated by more than 15A.     

Evidence for kinks in DNA folding in the nucleosome

The nucleosome subunit of chromatin consists of DNA folded around a histone core as a 1.8-turn left-handed solenoid. The crystal structure of the nucleosome core particle revealed that it has a dyad symmetry axis and that the minor helix groove faces outwards from the protein core. Richmond et al. noticed that the path traversed by the helix has severe bends at sites approximately one and four helix turns from the dyad axis. We have developed two photochemical methods to study the structure of DNA, and in particular that wrapped around the nucleosome core. One method depends on the sensitization of singlet oxygen production by an eosin analogue. We have monitored the rate at which excited state oxygen diffuses into contact with DNA base planes, and find that it attacks the nucleosome with high specificity. We have also mapped the DNA binding of the intercalating dye methylene blue, and conclude that it binds to the same sites accessible to oxygen by diffusion. On the basis of these results we suggest that the DNA in the nucleosome is bent or kinked at two sites, 1.5 helix turns from the dyad axis.     

Structural and Chemical Diversity of Tl－Based Cuprate Superconductors

The Tl-based cuprate superconductor family is the largest family in crystal structure and chemical composition among all high Tc cuprate superconductors. The Tl family can be divided into two sub-families, the Tl single layer family and the Tl double layer family, based on their crystal structural characteristics. The Tl single layer family is an ideal material for investigating the evolution of crystalline formation, charge carrier density, chemical composition, transport properties, superconductivity and their relationships. The Tl family contains almost all possible crystal structures discovered in high-Tc cuprate superconductors. Tl cuprate superconductors are of great importance not only in studying high-temperature superconductivity but also in commercial applications.     

Hierarchical self-assembly of DNA into symmetric supramolecular polyhedra

DNA is renowned for its double helix structure and the base pairing that enables the recognition and highly selective binding of complementary DNA strands. These features, and the ability to create DNA strands with any desired sequence of bases, have led to the use of DNA rationally to design various nanostructures and even execute molecular computations. Of the wide range of self-assembled DNA nanostructures reported, most are one- or two-dimensional. Examples of three-dimensional DNA structures include cubes, truncated octahedra, octohedra and tetrahedra, which are all comprised of many different DNA strands with unique sequences. When aiming for large structures, the need to synthesize large numbers (hundreds) of unique DNA strands poses a challenging design problem. Here, we demonstrate a simple solution to this problem: the design of basic DNA building units in such a way that many copies of identical units assemble into larger three-dimensional structures. We test this hierarchical self-assembly concept with DNA molecules that form three-point-star motifs, or tiles. By controlling the flexibility and concentration of the tiles, the one-pot assembly yields tetrahedra, dodecahedra or buckyballs that are tens of nanometres in size and comprised of four, twenty or sixty individual tiles, respectively. We expect that our assembly strategy can be adapted to allow the fabrication of a range of relatively complex three-dimensional structures.     

Crystal structure of an RNA double helix incorporating a track of non-Watson-Crick base pairs

The crystal structure of the RNA dodecamer duplex (r-GGACUUCGGUCC)2 has been determined. The dodecamers stack end-to-end in the crystal, simulating infinite A-form helices with only a break in the phosphodiester chain. These infinite helices are held together in the crystal by hydrogen bonding between ribose hydroxyl groups and a variety of donors and acceptors. The four noncomplementary nucleotides in the middle of the sequence did not form an internal loop, but rather a highly regular double-helix incorporating the non-Watson-Crick base pairs, G.U and U.C. This is the first direct observation of a U.C (or T.C) base pair in a crystal structure. The U.C pairs each form only a single base-base hydrogen bond, but are stabilized by a water molecule which bridges between the ring nitrogens and by four waters in the major groove which link the bases and phosphates. The lack of distortion introduced in the double helix by the U.C mismatch may explain its low efficiency of repair in DNA. The G.U wobble pair is also stabilized by a minor-groove water which bridges between the unpaired guanine amino and the ribose hydroxyl of the uracil. This structure emphasizes the importance of specific hydrogen bonding between not only the nucleotide bases, but also the ribose hydroxyls, phosphate oxygens and tightly bound waters in stabilization of the intramolecular and intermolecular structures of double helical RNA.     

Nature,nurture and human disease

What has been learnt about individual human biology and common diseases 50 years on from the discovery of the structure of DNA? Unfortunately the double helix has not, so far, revealed as much as one would have hoped. The primary reason is an inability to determine how nurture fits into the DNA paradigm. We argue here that the environment exerts its influence at the DNA level and so will need to be understood before the underlying causal factors of common human diseases can be fully recognized.     

Structural analysis of a eukaryotic sliding DNA clamp-clamp loader complex

Sliding clamps are ring-shaped proteins that encircle DNA and confer high processivity on DNA polymerases. Here we report the crystal structure of the five-protein clamp loader complex (replication factor-C, RFC) of the yeast Saccharomyces cerevisiae, bound to the sliding clamp (proliferating cell nuclear antigen, PCNA). Tight interfacial coordination of the ATP analogue ATP-gammaS by RFC results in a spiral arrangement of the ATPase domains of the clamp loader above the PCNA ring. Placement of a model for primed DNA within the central hole of PCNA reveals a striking correspondence between the RFC spiral and the grooves of the DNA double helix. This model, in which the clamp loader complex locks onto primed DNA in a screw-cap-like arrangement, provides a simple explanation for the process by which the engagement of primer-template junctions by the RFC:PCNA complex results in ATP hydrolysis and release of the sliding clamp on DNA.     

程序控温动态分析法对聚醚醚酮热稳定性的研究

以程序升温分解法和程序升温－原位红外谱法为研究手段，考察了热过程对ＰＥＥＫ结构特征的影响及其结构与热稳定性的关系。结果发现，在受热过程中，ＰＥＥＫ样品首先发生结晶结构的变化，不同红外吸收谱带对ＰＥＥＫ样品的结晶度敏感性不同；对不同的ＰＥＥＫ样品，由于它们在分子结构和结晶结构上存在差异，因此其热稳定性亦存在较大差异。     

Determination of the absolute handedness of knots and catenanes of DNA

DNA winds about itself in a right-handed or left-handed fashion at several structural levels. The double helix is generally right-handed and is given a (+) sign by convention, whereas supercoiling of the helix axis is always (-) in the cell. The winding in higher -order forms such as knots and catenanes is unknown, and this has impeded elucidation of the mechanisms of their formation and resolution by replication, recombination and topoisomerase action. We introduce here a procedure for determining the handedness of DNA winding by inspection of electron micrographs of DNA molecules coated with Escherichia coli RecA protein. We demonstrate the validity of the method and show that DNA topoisomerase I of E. coli generates an equal mixture of (+) and (-) duplex DNA knots, and that one product of recombination by resolvase of transposon Tn3 (refs 8, 9) is a catenane of uniquely (+) sign.     

反溶剂重结晶法制备多晶型比卡鲁胺

采用反溶剂重结晶法制备多晶型比卡鲁胺,通过改变重结晶过程中所使用的溶剂,对比单一溶剂和共溶剂对所制得晶体的晶型和颗粒形貌差别,探索最优化的共溶剂类型和体积比参数,以达到将比卡鲁胺原料药进行晶型纯化的目的。在反溶剂重结晶过程中采用共溶剂制备比卡鲁胺晶体发现：共溶剂（二甲基亚砜＋乙醇）以及（丙酮＋乙醇）可制备比卡鲁胺晶型I,并通过调节共溶剂中乙醇的用量可得到粒度均一,分散性好的I型比卡鲁胺。     

The three-dimensional structure of a DNA duplex containing looped-out bases

Unpaired bases in DNA have been assigned a possible role in the mechanism of frameshift mutagenesis in sequences with repeated base pairs. They also occur in quasipalindromic DNA sequences, which have been implicated in mutagenesis where there are no repeated base pairs, through the formation of single-stranded hairpin loops. The conformation of unpaired bases in DNA has been the subject of numerous thermodynamic as well as high resolution NMR (nuclear magnetic resonance) studies (reviewed in ref. 4). The NMR studies in solution have shown that the duplex of the tridecamer DNA fragment d(CGCAGAATTCGCG) remains intact, and that the unpaired adenosines are stacked into the duplex. Having crystallized this oligonucleotide and determined its structure, we find its conformation in the crystal is close to that of a B-DNA duplex, with the two additional adenosines looped out from the double helix and causing little disruption of the rest of the structure.     

DNA打开生命之门 —— 遗传、发育与进化

DNA分子双螺旋结构的揭示直接引导现代生命科学的基础理论——中心法则建立。在过去的短短的50年中,生命科学发生了“脱胎换骨”的变化,几乎所有的传统生命科学分支学科都获得了长足的进步,许多新兴的生物学分支学科和边缘学科相继建立,生物学技术引发了社会产业结构的深刻变革,许多生物学观念也已经渗透到社会生活的各个方面,它所打开的探索生命奥秘之门将继续引领人们去挑战更多的未解的生命之谜。