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Efficient procedures are described for the disassembly of Cowpea Chlorotic Mottle Virus (CCMV) into its viral-RNA and capsid-protein components, the separation of the RNA and protein, and the reassembly of the purified protein into higher order nanoscale structures. These straightforward biochemical techniques result in high yield quantities of protein suitable for further biophysical studies (AFM, X-ray scattering, NMR, osmotic stress experiments, protein phase-diagram) and nanotechnology applications (protein enclosed nanoparticles, protein-lipid nanoemulsion droplets). Also discussed are solution conditions that affect the stability of the self-assembled protein structure and explicitly show that divalent cation is not required to obtain stable protein structures, while the presence of even small amounts of Ba 2+ have a significant impact on protein self-assembly. However, since high ionic strength solution conditions result in good yields of CCMV-like protein capsids, it is suggested that the highly charged cationic protein N-terminus could act as an electrostatic switch for protein self-assembly and therefore be modulated by ionic strength and salt type. It was also found that CaCl2/RNA precipitation methods do not yield sufficiently pure protein samples. © 2007 Elsevier B.V. All rights reserved.

Ultrafiltration using polyethersulfone-membranes was evaluated as an efficient and preferred method for purifying Cowpea Chlorotic Mottle Virus (CCMV). Cesium chloride (CsCl) ultracentrifugation and ultrafiltration protocols are described, and comparative UV-spectroscopic and electron micrograph results are presented. CCMV purified by ultrafiltration are shown to be equivalent to CCMV purified by ultracentrifugation, while reducing purification time by two days and avoiding the need for expensive capital overheads such as ultracentrifuges, rotors and toxic CsCl chemical waste.

A novel nucleic acid secondary structure, exemplified by d(A+-G)lo, is formed by an intramolecular, cooperative, acid-induced, coil - helix transition. The helix is apparently left-handed, lacks base stacking and pairing, and is maintained by hydrogen and ionic bonds between dA+ "side- chain" residues (with electropositive hydrogens -N6H2, -N,+H) and the phosphodiester backbone. Modeling indicates that those dA+ residues lie approximately parallel to the helix axis, interacting with the n - 1 backbone phosphates (with electronegative oxygens), somewhat like the -C=O-H-N- longitudinal interactions in a protein a-helix. Moreover, the intervening dG side-chain residues are extrahelical, as are amino acid side chains of an a-helix. All currently recognized types of nucleic acid secondary structure involve helical stacks of bases. Such stacks occur in single strands, in base pairs of hairpins formed by single strands, in base pairs of two-stranded complexes, in base triplets of three-stranded structures, and in base quartets of four-stranded complexes. In this paper, we provide details of an entirely different type of nucleic acid helical secondary structure, one that does not depend for its stabilization on helically wound stacks of bases or base pairs. Rather, this novel type of secondary structure encompasses single strands of the repeating ho- mopurine doublet sequence d(A-G),, helically twisted by an unusual combination of ionic and hydrogen (H) bonds essentially parallel to the helix axis. We first came upon this nucleic acid secondary structure in the course of investigating the molecular properties of d(A- G)lo, which displays an intense differential absorption of circularly polarized light (CD) without concomitant develop- ment of hypochromism in the near-ultraviolet (UV) when the pH is reduced from neutrality. Additional observations enabled attribution of this unusual dichotomy in optical properties to a single-stranded helical secondary structure stabilized largely by Coulombic interactions between nega- tively charged backbone phosphates and distal protonated dA (dA+) residues that do not overlap with their nearest neighbor dG residues. Although these properties made it possible to deduce some regular features of the secondary structure of d(A+-G)lo, direct evidence for many structural ~

This paper addresses the general question: what are the significant guest properties selected by this host when interacting with guest molecules in the liquid phase, resulting in cocrystallization of the host and guest? In particular, to what extent do π electrons in a guest molecule effect its potential as a guest? Werner clathrates of the host [Ni(NCS)2(4-ViPy)4] with mixtures of tetrahydrofuran (THF) and cyclic hydrocarbons as guests have been synthesised and their structures elucidated. Clathrate (1): [Ni(NCS)2(4-ViPy)4](1.78 THF)(0.22 cyclohexane), crystallizes in the orthorhombic space groupPbcna=9.976(6),b=20.630(25),c=19.861 (4) A,V=4087A3,Z=4,R=0.087 for 1461 reflections; (2): [Ni(NCS)2(4-ViPy)4](1.76 THF)(0.24 cyclohexene),Pbcn,a=9.987(7),b=20.614(4),c=19.898(4)A,V=4096A3,Z=4,R=0.084 for 1304 reflections; (3): [Ni(NCS)2(4-ViPy)4](0.48 THF)(0.52 1,3-cyclohexadiene), tetragonalI41/a,a=16.898(3),b=16.898(3),c=26.463(6)A,V=7556A3,Z=8,R=0.120 for 1698 reflections; (4): [Ni(NCS)2(4-ViPy)4](0.36 THF)(1.04 1,4-cyclohexadiene),I41/a,a=16.986(4),b=16.986(4),c=25.896(15)A,V=7472A3,Z=8,R=0.103 for 2025 reflections; (5): [Ni(NCS)2(4-ViPy)4](0.35 THF)(1.05 benzene),I41/a,a=17.102(10),b=17.102(10),c=25.498(8)A,V=7458A3,Z=8,R=0.118 for 2200 reflections; (6): [Ni(NCS)2(4-ViPy)4](3 benzene), triclinicP1,a=10.432(24),b=11.155(9),c=21.581(7)A, α=78.70(5), β=82.60(7), γ=74.09(13)°,V=2361A3,Z=2,R=0.078 for 3427 reflections. Host-guest ratios and, for mixtures of guests, guest1/guest2 ratios, were elucidated by density and NMR. We show that the conformational freedom of the substituted pyridines is not the primary reason for the clathrating ability of Werner hosts. All six structures show no host-guest interaction at the level of van der Waals interactions. As non-bonding interactions are not observed between the host and guest, this study shows that the above host's selectivity by enclathration of particular guest molecules cannot be accounted for by solid state structural analysis.

The thermal stability of the triplexes d(C(+)-T)(6):d(A-G)(6);d(C-T)(6) and d(T)(21):d(A)(21);d(T)(21) was studied in the presence of high concentrations of the anions Cl(-), HPO(4)(2-), CH(3)COO(-), SO(4)(2-) and ClO(4)(-). Thermally-induced triplex and duplex transitions were identified by UV- and CD-spectroscopy and T(m) values were determined from melting profiles. A thermodynamic analysis of triplex transitions shows the limitations of commonly used treatments for determining the associated release or uptake of salt, solute or water. Enhancement of the stability of these triplexes follows the rank order of the Hofmeister series for anions of sodium and ammonium salts, whereas water structure-breaking solutes have the opposite effect. The rank order for the Hofmeister series ClO(4)(-)

Near-UV difference spectral analysis of the triplex formed from d(C-T)6 and d(A-G)6.d(C-T)6 in neutral and acidic solution shows that the third strand dC residues are protonated at pH 7.0, far above their intrinsic pKa. Additional support for ion-dipole interactions between the third strand dC residues and the G.C target base pairs comes from reduced positive dependence of triplet stability on ionic strength below 0.9 M Na+, inverse dependence above 0.9 M Na+ and strong positive dependence on hydrogen ion concentration. Molecular modeling (AMBER) of C:G.C and C+:G.C base triplets with the third strand base bound in the Hoogsteen geometry shows that only the C+:G.C triplet is energetically feasible. van't Hoff analysis of the melting of the triplex and target duplex shows that between pH 5.0 and 8.5 in 0.15 M NaCl/0.005 M MgCl2 the enthalpy of melting (delta H degree obs) varies from 5.7 to 6.6 kcal.mol-1 for the duplex in a duplex mixture and from 7.3 to 9.7 kcal.mol-1 for third strand dissociation in the triplex mixture. We have extended the condensation-screening theory of Manning to pH-dependent third strand binding. In this development we explicitly include the H+ contribution to the electrostatic free energy and obtain [formula: see text]. The number of protons released in the dissociation of the third strand from the target duplex at pH 7.0, delta n2, is thereby calculated to be 5.5, in good agreement with approximately six third strand dc residues per mole of triplex. This work shows that when third strand binding requires protonated residues that would otherwise be neutral, triplex formation and dissociation are mediated by proton uptake and release, i.e., a proton switch. As a by-product of this study, we have found that at low pH the Watson-Crick duplex d(A-G)6.d(C-T)6 undergoes a transition to a parallel Hoogsteen duplex d(A-G)6.d(C(+)-T)6.

Clark and White wonder why the chemistry education community has not uniformly adopted just one form of the periodic table. Part of the answer is that the majority who are silent on this issue do not want to be attacked by the vocal proponents who insist that lanthanum and actinium must be in the f-block and lutetium and lawrencium must be in the d-block.

This paper discusses the reasons and implications of placing lanthanum (La) and actinium (Ac) in the f-block and lutetium (Lu) and lawrencium (Lr) in the d-block. The author's conclusion is that placing lanthanum (La) and actinium (Ac) in the f-block results in a less satisfactory presentation of the elements in the periodic table and that lanthanum (La) and actinium (Ac) should remain in the d-block below yttrium (Y) in group 3 (IIIB).

The thermal characteristics of seven crystals with [Ni(NCS)2(vpy)4](vpy = 4-vinylpyridine) as host were analysed by thermal gravimetric analysis (TGA) and differential thermal analysis (DTA). No preferential guest loss for the clathrates with pairs of guests was observed. The enthalpy of the ‘guest release reaction’ for each clathrate was obtained from the DTA experiments. Solubility studies of the host were performed in the same guest solvents in order to obtain a better understanding of the clathration process. The results show that the basis of the selection of the host for guest molecules is related to solubility.

Bacterial viral capsids in aqueous solution can be opened in vitro by addition of their specific receptor proteins, with consequent full ejection of their genomes. We demonstrate that it is possible to control the extent of this ejection by varying the external osmotic pressure. In the particular case of bacteriophage λ, the ejection is 50% inhibited by osmotic pressures (of polyethylene glycol) comparable to those operative in the cytoplasm of host bacteria; it is completely suppressed by a pressure of 20 atmospheres. Furthermore, our experiments monitor directly a dramatic decrease of the stress inside the unopened phage capsid upon addition of polyvalent cations to the host solution, in agreement with many recent theories of DNA interactions.