Charged End Research Articles

Formation of Inclusion Complexes of Oligoethylene and Its Derivatives with α-Cyclodextrin

Abstract Oligoethylene (OE) was found to form inclusion complexes with α-cyclodextrin (α-CD) not only from aqueous solutions of α-CD but also from DMF solutions of α-CD to give stoichiometric compounds in a crystalline state in high yields. The yields depend on the oligomeric degree (n) of OE when DMF was used as solvent. OE of n less than 6 did not form complexes with α-CD in DMF solution. The complexes were characterized by IR, 1H NMR, 13C NMR, and 13C CP/MAS and PST/MAS NMR spectra, X-ray (powder), and elemental analysis. The continuous variation plots for the complex formation and the 1H NMR spectra of the complexes show that the stoichiometry of the crystalline complexes is 3 : 1 (ethylene unit–α-CD). The X-ray powder pattern of α-CD–OE complexes and the solid state 13C CP/MAS, PST/MAS NMR spectra suggest that the OE chain is included in the channel formed by α-CD molecules and that the OE backbone in the complexes is more flexible than that in uncomplexed state (OE crystal) due to being included in the cavities of α-CD. Derivatives of OE(6) with small end groups, such as hydroxyl, carboxyl and amino groups, formed crystalline complexes with α-CD, but OE(6) carrying large end groups, such as 2,4-dinitrophenyl and 2,4,6-trinitrophenyl groups, did not form complexes with α-CD. However, it is found that derivatives of OE(6) with charged end groups, such as ammonio or carboxylato groups, do not form crystalline complexes with α-CD, but can form 1 : 1 (guest molecule–α-CD) complexes which are in equilibrium in aqueous solution. The complexation of diammoniohexaethylene cation (DAHE cation) with α-CD in aqueous solution was discussed by using 1H NMR and 2D NOESY NMR spectra.

Amine blockers of the cytoplasmic mouth of sodium channels: a small structural change can abolish voltage dependence

Many drugs block sodium channels from the cytoplasmic end (Moczydlowski, E., A. Uehara, X, Guo, and J. Heiny. 1986. Isochannels and blocking modes of voltage-dependent sodium channels. Ann. N.Y. Acad. Sci. 479:269-292.). Lidocaine, applied to either side of the membrane, induces two blocking modes, a rapid, voltage-dependent open-channel block, and a block of the inactivated channel that occurs on a 1000-fold slower timescale. Here we describe the actions of several lidocaine-related amines on batrachotoxin(BTX)-activated bovine cardiac sodium channels incorporated into planar lipid bilayers. We applied blocking amines from the intracellular side and examined the structural determinants of fast, open-channel block. Neither hydroxyl nor carbonyl groups, present in the aryl-amine link of lidocaine, were necessary, indicating that hydrogen bonding between structures in the aryl-amine link and the channel is not required. Block, however, was significantly enhanced by addition of an aromatic ring, or by the lengthening of aliphatic side chains, suggesting that a hydrophobic domain strengthens binding while the amine group blocks the pore. For most blockers, depolarizing potentials enhanced block, with the charged amine group apparently traversing 45-60% of the transmembrane voltage. By contrast, block by phenylhydrazine was essentially voltage-independent. The relatively rigid planar structure of phenylhydrazine may prevent the charged amino end from entering the electric field when the aromatic ring is bound. The relation between structural features of different blockers and their sensitivity to voltage suggests that the transmembrane voltage drops completely over less than 5 A. We raise the possibility that the proposed hydrophobic binding domain overlaps the endogenous receptor for the inactivation gate. If so, our data place limits on the distance between this receptor and the intrapore site at which charged amines bind.

Dyeing Behaviors of Acid Azo Dyes with Alkyl Groups of Different Carbon Number in Nylon

Equilibrium sorption and diffusion rates of acid dyes in nylon dyeing were investigated. The dyes used have alkyl groups containing O-12 carbon atoms. The sorption isotherms were analysed quantitatively by a dual sorption model, consisting of partition (P) and Langumuir (L) mechanism. With increase in the number of carbon atoms, the P type contribution increases equilibrium sorption. Dyes containing alkyl groups with less than 10 of carbon atoms were sorbed stoichiometrically by the L type mechanism at the charged amino end groups of nylon. On the other hand, when the groups contained 10 and 12 of carbon atoms, the dyes were sorbed in excess of the equivalent of charged amino end groups.Diffusion profiles in the nylon substrate were determined by the cylindrical film roll method and the results were interpreted by the same dual model. The longer the alkyl group, the larger in the contribution of P type diffusion, but its contribution to total diffusion is small. The concentration dependence of the diffusion coefficient of the L type varied with the length of the alkyl chains.

Monte Carlo calculation of the conformational free energy and entropy of a polypeptide chain near a charged surface

The conformational behavior of a molecular model of enkephalin near a charged plane is studied using a Monte Carlo procedure. The variation of the dielectric permittivity near the charged surface is taken into account and the electric image method is used in order to obtain the interaction energy of the molecule with the plane as well as the electrostatic interactions between partial atomic charges. Evaluation of the free energy and entropy of conformation are made as functions of the molecule to plane distance. The molecular chain adopts folded structures near the surface and extended ones far from it, but the folding of the molecule and its attraction or repulsion by the plane depend largely on the charge density of that surface and on the charged ends of the molecule.

Static structure factor and shape of reptating telehelic ionomers in electric fields

The exact static structure factor of reptating block copolymers which have a neutral middle block and charged ends is calculated. In the presence of an electric field, these telehelic ionomers reptate randomly in their «tubes» but the latter tend to orient along the field axis. If the two ends have different charges, competition between many length scales occurs. The resulting scattering function shows unusual features that are normally characteristic of highly polydisperse mixtures. The results agree with a recent calculation of the radii of gyration for these molecules. The possibility of using electric fields to orient long polymers is discussed

Proteolytic modification of the heparin-binding affinity of extracellular superoxide dismutase.

The heparin-binding affinity of the tetrameric extracellular superoxide dismutase (EC-SOD) is a result of the cooperative effect of the heparin-binding domains of the subunits, located in the hydrophilic, strongly positively charged C-terminal ends. EC-SOD C, the high-heparin-affinity type, exposed to immobilized trypsin and plasmin was found to rapidly lose its affinity for heparin, without any loss of enzymic activity or major change in molecular mass as judged by size-exclusion chromatography. Heparin and dextran sulphate 5000 inhibited the proteolysis, suggesting that EC-SOD C sequestered by heparan sulphate proteoglycan in vivo is partially protected against proteolysis. The loss of heparin-affinity occurred with the stepwise formation of intermediates, and the pattern upon chromatography on heparin-Sepharose and subsequent immunoblotting was compatible with the notion that the changes are due to sequential truncations of heparin-binding domains from subunits composing the EC-SOD tetramers. A similar pattern with intermediates and apparent truncations has previously been found with EC-SOD of human plasma. The findings show that the unique design of the heparin-binding domain of EC-SOD allows easy modification of the heparin-affinity by means of limited proteolysis, and suggest that such proteolysis is a major contributor to the heterogeneity in heparin-affinity of EC-SOD in mammalian plasma.

Direct STEM fabrication and characterization of self‐supporting carbon structures for nanoelectronics

AbstractA quantitative model is proposed to interpret the experimental results concerning the observed anomalous behavior of the thickness of growing self‐supporting carbon‐containing rods: a decrease in the thickness with increasing beam current. The model is based on the surface diffusion of hydrocarbon molecules and takes into account their ionization and desorption from the positively charged end of the growing rod. Possible causes of self‐maintained oscillation instability of the rod growth are discussed. Procedures are recommended to optimize the fabrication of self‐supporting carbon‐containing structures of complicated configurations and to reduce the negative effect of the incident beam electrons during their testing.

Non-peptidic anti-aids agents: Inhibition of HIV-1 proteinase by disulfonates

Based upon an earlier observation that sodium docosanedioate (NaO 2C-(CH 2) 20-CO 2Na) weakly inhibits HIV-1 proteinase (IC 50 12μM), we have identified a class of more potent inhibitors (sulfonic acids) of this enzyme which are likewise dianionic at pH 5–6.5. Many of the compounds were moderately strong inhibitors of the enzyme (IC 50 40nM-10μM) and some have previously been shown to have anti-HIV activity in lymphocytes. Proteinase inhibition was dependent on the separation between sulfonate/carboxylate substituents, consistent with the hypothesis that negative charged ends of an inhibitor might form ionic bonds with Arg 8 and Arg 108 located at either end of the substrate-binding groove of the enzyme. The binding mode remains to be established by structure elucidation. Results for enzyme inhibition are presented along with structure-activity relationships and evidence for pH dependent inhibition. The general observations reported here may be useful for developing more potent and selective non-peptidic proteinase inhibitors.

The surface characterization of model charged and sterically stabilized polymer colloids by SSIMS and X.p.s.

X-ray photoelectron spectroscopy (X.p.s.) and static secondary ion mass spectrometry (SSIMS) have been applied to the surface chemical analysis of charged and sterically stabilized polymer colloids. The techniques provide complementary evidence for the surface orientation of both the charged polymer end groups and polymer chains responsible for the steric barrier. The findings are discussed in terms of the particle formation mechanisms and the biological interactions of such colloid systems.

Comparison of properties of conducting TCNQ salts with 1,4-bis[1-(4--r-pyridinio)]butane and trans-2-butene dications

Conducting TCNQ salts are prepared with dications consisting of butyl and trans-2-butenyl as the central linkages and various 4-substituted pyridinium as the charged end groups. Conductivities are comparable and of low values (10−4 − 10−2 S/cm) for the salts having H, t-butyl and benzyl as the substituents of pyridine irrespective of central linkages. For the salts with 4-methyl and 4-ethyl as the substituents of pyridine, the conductivities are 4 orders of magnitude higher for butyl linkage. UV-VIS-Near IR and IR spectra of these salts are presented. The variation of conductivity is explained in terms of dication structures.

Inhibition of HIV-1 proteinase by non-peptide carboxylates

Some simple dicarboxylates are among the first reported non-peptide inhibitors of HIV-1 proteinase. Only weak inhibition (IC 50 ≥ 10μM) was observed but this may be significant since only two potential enzyme-binding groups are present. Dixon plots and preliminary kinetic data are reported and a possible mechanism for the inhibition is discussed. The dicarboxylates are long enough to engage the carboxylate side chains of Arg 8 and Arg 108 at either end of the 24Å long substrate-binding groove. This mode of binding has not been proven but other molecules with similarly separated charged ends are equally effective inhibitors, perhaps indicating a common mechanism of inhibition. There is evidence that placing other functional groups on the inhibitor enables alternative interactions with the enzyme which can reduce inhibitor potency. We propose that incorporation of ionic binding groups in more elaborate and selective non-peptides may potentiate inhibition of HIV-1 proteinase.

Photophysical and photochemical properties of rhodamine 6G in alcoholic and aqueous sodium dodecylsulphate micellar solutions

The photodecomposition of the rhodamine 6G (R6G) dye dissolved in n-alcohols and aqueous sodium dodecylsulphate (SDS) micellar solution was investigated under continuous UV-visible illumination. The results indicate that in alcohol solution the photodecomposition efficiency increases with the dye concentration. It also depends on the nature of the alcohol the lowest efficiency being observed in ethyleneglycol. The mechanism involves the electron transfer reaction of the triplet excited state with a ground state dye molecule (T + S 0 → R + + R − . In aqueous SDS micellar solution, the R6G cation is embedded in the hydrophobic interior of the micelle close to the negatively charged sulphate end groups. We show that a rather high concentration (3 × 10 −3 M) of R6G can be dissolved in 10 −1 M SDS aqueous solution, protecting R6G from the electron transfer reaction and secondary decomposition reactions.

A negatively charged N terminus in the alpha polypeptide inhibits formation of light-harvesting complex I in Rhodobacter capsulatus

Light-harvesting complex I (LHI) of Rhodobacter capsulatus contains bacteriochlorophyll and carotenoids which are noncovalently bound to two different apoproteins (alpha and beta polypeptides) carrying oppositely charged N-terminal ends. The contribution of these charged segments to the assembly of LHI was studied with mutants having oppositely charged amino acids in the alpha or beta polypeptide. The influence of these mutations on the insertion and assembly process of the LHI complex was investigated by means of spectroscopic analysis of isolated intracytoplasmic membranes and pulse-chase experiments. Exchange of four positively charged amino acids to negatively charged amino acids on the N-terminal domain of the alpha subunit inhibited completely the assembly of the LHI complex. Although this mutant has no antenna, the reaction center is active and the cells were able to grow anaerobically in the light. Conversely, mutation of the four negatively charged amino acids of the N-terminal segment of the beta polypeptide did not prevent the assembly of the LHI complex, although the stability of the complex and the size of the photosynthetic unit were affected. The presence of the mutated beta polypeptide was confirmed by protein sequencing.

Electrostatic potential mapping of polycyclic aromatic hydrocarbon diol epoxides using a dipole

AbstractElectrostatic potential maps of benz(a)anthracene diol epoxide, benzphenanthrene diol epoxide, chrysene diol epoxide, and tryphenylene diol epoxide, which are ultimate carcinogens derived from the corresponding polycyclic aromatic hydrocarbons, have been studied using a dipole of length 1 Å and strength 1 Debye. The net charge distributions in the molecules were obtained using the MNDO molecular orbital method. The maps were drawn for two closest distances of approach between the charged ends of the dipole and atoms of the molecules. Using the electric field directions and values of electrostatic potentials, reactive sites and relative reactivities of these molecules have been examined. Existence of a bay region appears to be important for the carcinogenic activity of a diol epoxide in this class of carcinogens.

Role of chain termini in selective steroidogenic effect of ACTH/MSH(4–10) on isolated adrenocortical cells

To investigate the role of charged chain ends in the corticosteroidogenic effect of ACTH/MSH(4–10), acetyl and amide derivatives of ACTH/MSH(4–10) were synthesized and tested in isolated zona glomerulosa and zona fasciculata cells. ACTH/MSH(4–10)-NH 2, Ac-ACTH/MSH(4–10) and Ac-ACTH/MSH(4–10)-NH 2 (10 μM to 1 mM) stimulated the aldosterone production of zona glomerulosa cells, whereas these peptides did not stimulate the corticosterone production of zona fasciculata cells, even at 1 mM concentration. As ACTH/MSH(4–10) has been shown to have a steroidogenic effect on both types of adrenocortical cells, both charged chain termini seem to be essential for triggering of the corticosterone production of zona fasciculata cells, but for aldosterone production their presence appears not to be important.

Stabilization of protein structure by interaction of alpha-helix dipole with a charged side chain.

The alpha-helix in proteins has a dipole moment resulting from the alignment of dipoles of the peptide bond which can perturb the pKas of ionizing groups. One of the two histidine residues (His18) in barnase, the small ribonuclease from Bacillus amyloliquefaciens, is located at the negatively charged end (C-terminal) of an alpha-helix. From NMR titrations of wild-type and engineered mutants we find that the pKa of His18 is 7.9 in wild-type enzyme, 1.6 units above the value in the urea-denatured enzyme and in model peptides. This implies that there is a favourable interaction between the protonated form of His18 and the alpha-helix that should stabilize the native structure at neutral pH by 2.1 kcal mol-1. Denaturation at various values of pH of wild-type and muant enzymes engineered at position 18 shows that this is so. The increase in stability of the enzyme as the pH changes from 8.5 to 6.3 is attributable to this interaction, and the pH-stability curve fits pKa values for His18 in native and urea-denatured enzymes that are consistent with the NMR data.

Interaction of synthetic analogues of distamycin and netropsin with nucleic acids. Does curvature of ligand play a role in distamycin-DNA interactions?

Distamycin and netropsin, a class of minor groove binding nonintercalating agents, are characterized by their B-DNA and A-T base-specific interactions. To understand the conformational and chemical basis of the above specificities, the DNA-binding characteristics of a novel synthetic analogue of distamycin have been studied. The analogue, mPD derivative, has the requisite charged end groups and a number of potential hydrogen-bonding loci equal to those of distamycin. The difference in the backbone curvatures of the ligands, distamycin, the mPD derivative, and NSC 101327 (another structurally analogous compound), is a major difference between these ligands. UV and CD spectroscopic studies reported here show the following salient features: The mPD derivative recognizes only B-DNA, to which it binds via the minor groove. On the other hand, unlike distamycin, it binds with comparable affinities to A-T and G-C base pairs in a natural DNA. These DNA-binding properties are compared with those reported earlier for distamycin and NSC 101327 [Zimmer, Ch., & Wahnert, U. (1986) Prog. Biophys. Mol. Biol. 47, 31-112]. The backbone structures of these three ligands were compared to show the progressive decrease in curvatures in the order distamycin, mPD derivative, and NSC 101327. The plausible significance of the backbone curvature vis-à-vis the characteristic B-DNA and AT-specific binding of distamycin is discussed. To our knowledge, this is the first attempt (with a model synthetic analogue) to probe the possible influence of backbone curvature upon the specificity of interactions of the distamycin class of groove-binding ligands with DNA.

A Monte Carlo method with different treatments for short- and long-range interactions in conformational statistics of polypeptide chains

Separation of the conformational energy into terms associated with short-range or long-range interactions allows us to propose a new approach for using Monte Carlo calculations in statistical mechanics of molecular chains. The present method is based on a simultaneous use of statistical weight matrices calculated with short-range interactions and importance sampling applied only on the part of energy coming from long-range interactions. Distributions of end-to-end distances and probabilities of the chain units being in their different conformational states are determined on molecular models of the peptidic hormones enkephalin and β-casomorphin. Results obtained in this way are compared with values calculated using previous computational methods which are also applied on chain models including only nearest neighbor interdependent units. The relative importances of short-range and long-range interactions for the molecular conformation can thus be distinguished. Effects of residue Pro as well as of charged ends on the chain folding are well verified. It is shown that solvent effects can be introduced on long-range interactions mainly by screening the electrostatic term of the energy; extended conformations are then obtained. The proposed method lowers the computer time and by using more information on the moledular system, improves Monte Carlo calculations.

Cloning, expression, and primary structure of a Chlamydia trachomatis binding protein.

The gene encoding an 18,000-dalton eucaryotic cell-binding protein of Chlamydia trachomatis serovar L2 was cloned into Escherichia coli, and the nucleotide sequence of a 1,658-base-pair PstI restriction endonuclease fragment encoding this protein was determined. The recombinant chlamydial gene consists of a 486-base-pair open reading frame encoding a polypeptide of molecular weight 18,314. The resultant polypeptide, comprising 162 amino acids, possesses a highly charged carboxy-terminal end. The expression of this recombinant protein is under the control of a vector promoter. The recombinant 18,000-dalton protein possessed the same eucaryotic cell-binding characteristics as did the native chlamydial 18,000-dalton protein when electrophoresed and transferred to nitrocellulose. Polyclonal antibodies to the recombinant protein exhibited neutralizing activity.

Conformation of arginine side chain in a mini peptide. Molecular structure of acetyl‐l‐arginine‐ethylamide

The crystal structure of N‐α‐acetyl‐l‐arginine ethylamide perchlorate (molecular formula: C10H21N5O2 ± HClO4) has been determined from X‐ray diffraction data. This arginine derivative has the two charged ends substituted by peptide units and thus becomes representative of an arginyl residue in a protein. The crystals are orthorhombic, space group P21,21,21, a = 7.485 (2), b = 28.623 (5), c = 7.261 (3) Å, Z = 4. The structure was solved by direct methods and refined by full matrix least‐squares using 559 reflections to an R = 0.052. The molecule shows two planar peptide units with φ and ø values in the β pleated sheet region. The side chain torsion angles are in trans conformation, except the χ1, angle which is in the gauche(‐) region. This observation further illustrates that this is one of the favoured side chain conformations of arginine, together with the all‐trans conformation. Symmetric double hydrogen bonds are observed between the guanidinium group and perchlorate anions. The H(N) and oxygen atoms of the amide groups also form hydrogen bonds.