Right-handed Helical Structure Research Articles

NMR studies of an oligonucleotide with an unusual structure induced by platinum anticancer drugs

The 31P NMR spectra of Pt( en){ d( T 1 A 2 T 3 G 4 G5 G 6 T 7 A 8 C 9 C 10 C 11 A 12 T 13 A 14)} (14-me r) and Pt( en){ d( A 2 T 3 G 4 G 5 G 6 T 7 A 8 C 9 C 10 C 11 A 12 T 13)} (12-mer) (en = ethylenediamine) each contai far downfield ( ca. −2.9 and −2.6 ppm from trimethylphosphate standard), two signals slightly downfield, and at least one signal slightly upfield of the normal range ( ca. −4.0 to −4.4 ppm). This pattern suggested a distorted structure. The unusual 31P signals of the 12-mer were assigned by analogy to signals of the 14-mer previously assigned by 17O-labeling methods. A combination of heteronuclear multiple-quantum coherence, one-dimensional and two-dimensional nuclear Overhauser effect (1D- and 2D-NOE) and homonuclear shift correlation spectroscopy (COSY) experiments assigned all aromatic 1H signals of the 12-mer except H8 of G 5 or G 6. One of these H8 signals is missing from the spectrum and the nucleotide is labeled G m. The other H8 is the most downfield signal and has a strong NOE to its H1'. Since this strong NOE indicates that this nucleoside exists in a syn conformation, it is labeled G s. A strong NOE was observed between the G s and A 8 H8 signals. Several lines of evidence suggested a hairpin-like structure with a loop region (G 6T 7A 8C 9) and a stem region involving A 2T 3G 4G 5 and C 10C 11A 12T 13. The 31P signals for the stem region are within or slightly outside the normal range, 3 J H3'-Pvalues (3–6 Hz), measured by a 2D- J experiment, of stem nucleotides were characteristic for a DNA duplex. Imino signals for base pairs A 2T 13,T 3A 12, G 4C 11, and probably G 5C 10, and the observation of internucleotide NOE connectivities for these nucleotides (e.g. between an H8 signal and the H1' signal of the 5' nucleotide) suggested a right-handed helical structure. For the loop region, a distorted sugar-phosphate backbone is indicated by far downfield positions of the G 5pG 6 and A 8pC 9 31P signals, the if 3J H3′−P values for C 9p (8.0 Hz) and A 8p (6.8 Hz), and the absence of H3'-P coupling for G 5p. In the loop region, no imino signals or internucleotide NOEs characteristic of a right-handed duplex were observed. However, A 8H8, C 9H6, and C 10H6 each exhibited unusual internucleotide NOEs to the H4' signal of the 5' residue. NOE crosspeaks between T 7 1H signals and signals attributed to sugars of the G s and G m suggested that the T 7 moiety is located within the space encircled by the loop. The few NOE crosspeaks, pH dependence, and Cu 2+ broadening of C 9 1H signals indicate an isolated location accessible to solvent. Selective reverse chemical shift correlation, 1D- and 2D-NOE, and COSY experiments for the 14-mer produced results very similar to those described above for the 12-mer. We evaluated the following four models for the structure of the loop region of the 12-mer and the 14-mer: (1) Model I, in which Pt is bound to G 5G 6; (2) Model II, in which Pt is bound to three bases, G 5, G 6, and A 8; (3) a variation of Model II (Model IIC), in which one of the G bases forms a Pt—C8 bond; and (4) Model III, in which Pt is bound to A 8 and either G 5 or G 6. Model I does not explain the NOE between the A 8 and G s H8 signals nor the missing H8 signal. Two sets of results provide evidence against Model II and Model IIC. First, the 195Pt NMR spectra of the 12-mer and 14-mer have signals (−2645 and −2640 ppm respectively) similar to the reported shift for (Pt(en)(Guo) 2-N7,N7) 2+. Second, the presence of all fifteen signals in the 9–7 ppm region for 12-mer after Pt removal by KCN in D 2O suggested that Pt does not bind at C8. Model III provides the best explanation for our results. A pH dependence study of the 12-mer tentatively assigned G s as G 5. In summary, we believe that Model III, in which Pt binds G 5, is most reasonable and that a bend may occur at the intersection of the loop and stem regions.

Enhanced recA Protein Binding to Z DNA Represents a Kinetic Perturbation of a General Duplex DNA Binding Pathway

recA protein binding to duplex DNA is enhanced when a B form DNA substrate is replaced with a left-handed Z form helix. This represents a kinetic rather than an equilibrium effect. Binding to Z DNA is much faster than binding to B DNA. In other respects, binding to the two DNA forms is quite similar. recA protein binds to B or Z DNA with a stoichiometry of 1 monomer/4 base pairs. The final protein filament exhibits a right-handed helical structure when either B or Z form DNAs are bound. There are only two evident differences: the kcat for ATP hydrolysis is reduced 3-4-fold when Z DNA is bound, and recA binding at equilibrium is less stable on Z DNA than on B DNA. At steady state, the binding favors B DNA in competition experiments. The results indicate that Z DNA binding by recA protein follows the same pathway as for recA binding to B DNA, but that the nucleation step is faster on the Z form helix.

Thermotropic Cellulose Derivatives with Flexible Substituents. I. Preparation of Tri-o-(β-methoxyethoxy)ethyl Cellulose and its Cholesteric Mesophase Properties

Abstract Tri-o-(β-methoxyethoxy)ethyl cellulose was prepared by reacting the regenerated cellulose with 2-(2-methoxyethoxy)ethyl iodide and characterized as a fully substituted derivative. The resultant derivative forms a thermotropic liquid crystal in the wide temperature range from room temperature to the isotropization temperature of 180°C, showing that the (methoxyethoxy)ethyl side-chain substituent is long and flexible enough to inhibit crystallization of the extended cellulose chains and facilitate formation of liquid crystals. This is the first example of a thermotropic liquid crystalline cellulose derivative with a homogeneous side chain. This type of liquid crystal is cholesteric with a right-handed helical structure. The cholesteric pitch increases with increasing temperature, with the magnitudes comparable to the wavelength of visible light in the temperature range below 120°C.

核酸の分子構造及び核酸・蛋白質相互作用機構についてのX線解析法による構造研究

Since the discovery of the right-handed double helical structure of deoxyribonucleic acid by Watson and Crick in 1953, many pieces of detailed nucleic acid structural information involving ribonucleic acid have been elucidated and this review describes the results of the X-ray structural studies of nucleic acids and their constituents carried out in my laboratory. In the latter half the nucleic acid-protein interaction mechanism was also discussed on the basis of the molecular model of the stacking interaction between base and aromatic amino acid side chains. The structure-function relationship of ribonuclease T1.guanosine monophosphate complex was used as an example.

Mean geometry of the thiopeptide unit and conformational features of dithiopeptides and polythiopeptides

The mean geometry of the thiopeptide [C aNC(S)C a] unit has been derived from an analysis of X-ray crystal structure data, as well as MM2 and Gaussian 80 82 calculations. The conformational flexibilities of dithiopeptides with glycyl- and alanyl-side chains have been investigated by molecular mechanics. Minimum energy conformations were examined using interactive computer graphics molecular modeling techniques. Alanyl-dithiopeptide substitution within an oligopeptide results in considerable restriction of conformational freedom whereas the effect is minimal for glycyl-dithiopeptide substitution. Polyglycyl-thiopeptide adopts a left-handed three or fourfold or right-handed threefold helical structure with favorable interchain CS…HN hydrogen bond interactions. A poly-L-alanyl-thiopeptide prefers a left-handed threefold poly-L-proline-like helical structure.

Helical structure of β(1–3) xylan and the water mediated hydrogen bonding schemes

A six-fold intertwined triple helical structure for the polysaccharide β(1–3) xylan was generated with the axial advance of 0·306 nm per residue. A stereochemically possible site for the water molecule has been determined and water mediated intrachain and interchain hydrogen bond schemes are possible for the right-handed triple helical structure, whereas only interchain hydrogen bonding appears plausible in the left-handed triple helical structure. The water mediated hydrogen bond is almost linear. X-ray refinement using a Linked-Atom Least-Squares (LALS) procedure has enabled us to determine the orientation of the molecule in the hexagonal unit cell, locate the position of the water molecules and yield a reliability index, R, of 0·35. The refined model in this present study confirms the original chirality of an earlier model but differs in the water mediated hydrogen bonding scheme.

Conformation of ribooligonucleotide duplexes containing an alternating C-G sequence which show an unusual circular dichroism spectrum.

The poly[r(G-C)] duplex shows an unusually large negative band in the long wavelength region of the CD spectrum. In order to elucidate this phenomenon, r(C-G-C-G) and r(C-G-C-G-C-G) were synthesized chemically and their properties were examined by UV and CD, and 1H and 31P NMR spectroscopy. These ribooligomers form a self-complementary duplex at low temperature, the CD spectrum of which shows a negative band at around 290 nm and a positive band at around 265 nm with almost equal magnitudes. The proton resonances in the 1H NMR spectra of the oligo[r(C-G)] duplexes were assigned by nuclear Overhauser effect experiments. The chemical shift-temperature profiles of the base proton signals and the sharp singlets observed for all H1' protons are consistent with a normal A-RNA structure but not with a Z-DNA like structure. Moreover, a 500-MHz two-dimensional nuclear Overhauser effect experiment recorded for r(C-G-C-G-C-G) shows that all guanine bases adopt the normal anti-conformation. CD-temperature profiles and 31P NMR spectra of oligo[r(C-G)]s support this conclusion. These results indicate that duplexes of oligo- and polyribonucleotides containing alternating C-G sequences can give an unusually large negative CD band in the long wavelength region despite their right-handed helical structure.

Self base pairing in a complementary deoxydinucleoside monophosphate duplex: crystal and molecular structure of deoxycytidylyl-(3'-5')-deoxyguanosine.

The molecular structure of ammonium deoxycytidylyl-(3'-5')-deoxyguanosine, crystallized from aqueous acetone near pH 4, was determined for X-ray diffraction data. The crystals were tetragonal, space group P43212 with a = b = 11.078 (1) A and c = 45.826 (4) A. The structure was solved by tangent expansion of phases based on a derived phosphorus position and refined to R = 0.060 by full matrix least squares. Molecules related by a 2-fold symmetry axis are connected by hydrogen bonds between the bases and form parallel right-handed duplexes. Pairs of cytosines share a proton at N(3) and are joined by three hydrogen bonds: N(4)-H...O(2)...H-N(4), and N(3)-H...N(3). Guanines are joined by two hydrogen bonds: N(2)-H...N(3) and N(3)...H-N(2). Base-stacking interactions within the duplex are weak with the cytosine and guanine ring planes inclined at 24 degrees to each other in each monomer. Despite the unusual arrangement of the molecules, the sugar phosphate backbone has the g-g- conformation normally associated with right-handed double helical structures. Conformational parameters of the nucleosides are also typical with both sugars C(2')-endo and glycosidic torsion angles 55 degrees for cytidine and 94 degrees for guanosine. The bonding geometry of the bases is influenced by hydrogen bonding and charge-transfer networks in the crystal lattice. The solvent molecules interact with the dimer in three fused circular hydrogen bonding domains with a single disordered ammonium cation per d(CpG) dimer. Parallels with the formation of self base pairs and their implications in molecular biology are discussed.

Role of non-planar peptide unit in regular polypeptide helices. New model for poly-beta-benzyl-L-Aspartate.

The effect of non-planarity of the peptide unit on helical structures stabilized by intrachain hydrogen bonds is discussed. While the present calculations generally agree with those already reported in the literature for right-handed helical structures, it is found that the most stable left-handed structure is a novel helix, called the delta-helix. Its helical parameters are close to these reported for poly-beta-benzyl-L -aspartate. Conformational energy calculations show that poly-beta-benzyl-L -aspartate with the delta-helical structure is considerably more stable than the structure it is generally believed to take up (the omega-helix) by about 15 kcal/mol-residue.

Formation of helical filaments by copolymerization of two types of 'straight' flagellins.

Bacterial flagella undergo transition between several discrete types of left-handed and right-handed helical structures when exposed to acidic or alkaline pH, or to mechanical force. Calladine and ourselves have presented models to explain such polymorphism, assuming that protein subunits (flagellin) in a flagellum can be transformed into two conformations (L- and R-states) depending on the species of flagellin and on the environmental conditions. An obvious prediction from these 'two-state' models is that there should be two types of straight flagella (L- and R-types) that are made up exclusively of flagellins in either the L-state or the R-state. We have shown that straight flagella from two species of mutants, Salmonella SJ814 (ref. 6) and Escherichia coli hag 177 (ref. 7), are closely similar to the predicted R- and L-types, respectively. Recently we have isolated 10 strains of straight-flagellar mutants of Salmonella. We show here that their flagella can also be classified into the L- and R-types, and that copolymerization of flagellins from two heterologous types (L and R) makes discrete types of helical filaments, whereas that of homologous pairs of flagellins (L and L, or R and R) makes only straight filaments.

Helical Sense and Pitch of Cholesteric Liquid Crystals

Abstract The rotatory sense of a circularly polarized light transmitted through a cholesteric liquid crystal cell was studied by measuring the retardation of a birefringent plate placed upon the cell. Furthermore, the dependence of helical pitch on composition is described for various binary cholesteric and nematic-cholesteric mixtures. The helical sense of eleven cholesterics was determined by the experimental results. As a result, it is shown, for example, that cholesteryl chloride has a right-handed helical structure and cholesteryl propionate has a left-handed helical structure.

Structures of enzyme-substrate complexes of lysozyme

Conformational energy calculations were used to determine the binding structures of two oligosaccharides (GlcNAc)(n), in which n = 5 and 6, in the rigid active site of lysozyme (mucopeptide N-acetylmuramoylhydrolase, EC 3.2.1.17). Starting with the lowest energy binding structures of (GlcNAc)(4) as determined in a previous publication, we added a fifth GlcNAc residue to this tetramer in three different conformations, corresponding to the left-handed and right-handed helical structures and an intermediate structure, and the energy of each complex was minimized. The most stable binding conformation of the fifth residue of the pentamer was closest to the left-handed helical one. During energy minimization, the fourth residue of the pentamer moved from its initial position near the surface of the active site farther into the active site cleft at binding site D. Binding structures of (GlcNAc)(6) were then examined by addition of a residue to the lowest energy structure of (GlcNAc)(5), and it was found that the sixth residue of the hexamer binds in a conformation again close to the left-handed helical one. Stable binding regions of the rigid active site for the fifth and sixth residues were found to be near arginyl 45 and asparaginyl 46, on the opposite side of the active site cleft from arginyl 114. When the calculated structure of the lysozyme-(GlcNAc)(4) complex (used here as the starting structure for addition of the fifth and sixth residues) is compared with recent experimental data, it is found that the calculated structure is a reasonable one. Of all binding regions available to the saccharide residues, the C site binds GlcNAc with the lowest energy, in agreement with experiments.

Biological significance of helical conformation of acid polysaccharides.

Optical rotary dispersion spectra have been studied of methylene blue complexes of glycosaminoglycans and different derivates of chondroitin 4-sulphate (free and protein bound complexes of different molecular weights and degrees of sulphation). Structural requirements of Ch-4S induction of the extrinsic Cotton effect in methylene blue were formulated. This Cotton effect is supposed to reflect the right-handed helical structure of the polysaccharide molecules. Ch-4S displayed induced Cotton effect both in free and in protein bound form; molecular weight should exceed 2-3000 (at least 4-6 disaccharide units), degree of sulphation should exceed 3% sulphate-sulphur content (more than 0.5 sulphate per one disaccharide unit). For comparison the effect of different derivatives of Ch-4S on the rate of in vitro collagen fibril formation was investigated. Close correlation was found between the macrostructural characteristics and the possible biological effect of Ch-4S. The biological significance of the macrostructural properties of Ch-4S is discussed.

Dissymmetric recognition of the helical sense of deoxyribonucleic acid and evidence for binding of reporter molecules from the minor groove of DNA

Through the utilization of optically active DNP-derivatives of l- and d-proline, evidence is presented which suggests that nucleic acids exist as right-handed helices in solution. The results of ultraviolet absorption, circular dichroism, proton magnetic resonance (pmr), T m of the helix-coil transition, viscometric, and binding studies are consistent with the above interpretation. It is shown that several types of DNA (i.e., salmon sperm, calf thymus, Micrococcus luteus, poly d(A-T)-poly d(A-T) and poly d(I-C)-poly d(I-C)) exist in a right-handed helical structure in solution. In addition, evidence is presented which strongly indicates that the 2,4-dinitroaniline ring of DNP-proline is intercalated between base-pairs of DNA and the prolyl side chain situated in the minor groove. Moreover, it is shown that the more sterically hindered DNP-derivatives exhibit a higher selectivity for A-T binding sites.

Bacteriophage T2 as seen with the freeze-etching technique

Freeze-etching of bacteriophage T2 reveals new data on the phage ultrastructure which are discussed in relation to information obtained from negatively stained preparations. The polygonal head of the freeze-etched phage shows capsomeres of 10 nm (= 100 Å) in diameter with central depressions. There are six neighbors around most of the capsomeres, and the repeat vector of the surface lattice measures about 10 nm. Individual capsomeres seem to be composed of smaller units. The sheath of the phage tail exhibits a surface lattice indicative of six right-handed helices. The helices reveal a beaded fine structure, the elements of which are interpreted as homologous to the subunits seen after negative staining. The neighboring helices are cross-linked by bridgelike elements. During sheath contraction the pitch angle of the helices increases from 50 ° to 66 °. In the contracted sheath twelve helices are visible, again composed of subunits and cross-linked to their neighbors. The surface structure of the “polysheath” resembles that of a contracted sheath. Polysheaths break down into right-handed helical structures.

A second right-handed helical structure with the parameters of the Pauling-Corey alpha-helix.

Stereochemical studies predict bent hydrogen bonding between some CO and NH groups in the peptide chains of α-helices. X-ray crystallography has revealed the existence of such bonds in myoglobin and lysozyme.