Side Chain Density Research Articles

Entropy, likelihood and phase determination.

Entropy, likelihood and phase determination.

The Refined Crystal Structure of Hexon, the Major Coat Protein of Adenovirus Type 2, at 2·9 Å Resolution

The crystal structure of hexon, the major coat protein from adenovirus type 2, has been refined at 2·9 Å resolution. Hexon is a homo-trimer (molecular mass 3×109,077 Da) and crystallizes in the cubic space group P 21 3, with a cell edge of 150·5 Å. There are four molecules in the unit cell so that the crystallographic asymmetric unit contains one subunit of the trimer. The electron density in most regions is well-defined and 880 amino acid residues, of the 967 in this unusually long polypeptide chain, have been located and fitted. The N terminus (1 to 43) and three internal stretches (192 to 203, 270 to 291 and 444 to 453) are not defined, and a stretch (168 to 207) with unclear side-chain density is modelled as poly(Ala/Gly). The current refined model, consisting of 6943 non-hydrogen protein atoms and 85 water molecules, yields an R-factor of 19·9% for 18,176 reflections in the resolution range 5·0 to 2·9 Å. The model has reasonable geometry with root-mean-square deviations from ideal bond lengths of 0·022 Å and angle-related 1-3 distances of 0·056 Å.The overall shape of the trimeric hexon molecule is unusual and may be divided into a pseudo-hexagonal base rich in β-structure, and a triangular top formed from three long loops containing some secondary structure. The base contains two similar pedestal domains, P1 and P2, each of which is a flattened eight-stranded β-barrel with the "jelly-roll greek key" topology characteristic of other viral coat proteins. P1 and P2 are related by an approximate 6-fold operation about the molecular 3-fold axis so that six barrels form the walls of the tubular hexon base. The hexon bases form close-packed p3 arrays on each facet of the icosahedral adenovirus virion. Unlike other vital capsids, the barrel axes are almost perpendicular to rather than parallel with the capsid surface. The hexon top, which consists of intimately interacting loops emerging from P1 and P2 in the base, has a triangular outline and so does not exhibit the pseudo-symmetry of the base.The structure of the hexon trimer shows how economically it meets the demands of its function as a stable protective viral coat, reveals the significance of the special features in its unusual amino acid sequence, and explains its biochemical and immunological properties. The molecule is hollow, with a large central cavity, and so has a high effective volume for its mass. Hydrophobic residues are clustered together in the interior of P1 and P2, and at the central depression in the top of the molecule. The latter is a "hydrophobic plug" that seals the central cavity and so shields the viral interior from the outside environment. Intra and intersubunit salt bridges, hydrogen bonds and hydrophobic contacts all contribute to the extreme stability of hexon. In the trimer, each hexon subunit is locked in position to confer additional stability. The interaction of the loops at the top mad the N-terminal arm at the bottom prevents any relative upward or downward displacement of a subunit with respect to its neighbors. The intricate topology of the subunits suggests why their correct folding into the trimeric molecule requires the initial formation of a complex of hexon polypeptides with the adenovirus 100 K protein.

Ultrathin self-assembled polymeric films on solid surfaces. III. Influence of acrylate dithioalkyl side chain length on polymeric monolayer formation on gold

Self-assembled films of acrylate polymers containing dithioalkyl side chains of varying lengths have been fabricated on gold substrates by adsorption from dilute organic solution. Anchoring alkyl side chain types studied include lipoate (n=4), pentyl dithioundecanoate (n=10), pentyl dithiopalmitate (n=15), and pentyl dithiotricosonate (n=22), where n represents the number of methylene units in the longer arm of the asymmetric disulfide side chain. Comprehensive characterization of polymer monolayers by x-ray photoelectron spectroscopy and reflection Fourier transform infrared spectroscopy showed improved order for structural assemblies of C11 (n=10) side chain polymer monolayers, over shorter and longer side chain polymer analogs, due to a higher percentage of bound thiolate anchors. Monolayer thicknesses range from 20 to 40 Å, primarily depending on side chain length and density along the polymer backbone. Cyclic voltammetry on gold electrodes shows that longer side chain polymer monolayers possess more structural defects resulting from considerable disorder in the films. Despite the less organized structural features for these polymer monolayers, their selective adsorption onto gold via specific side chain terminal disulfide anchors on microlithographed substrate patterns creates well-resolved surface-modified microstructures comparable to those from monomeric analogs, as shown by scanning Auger mapping.

X-ray investigations of Langmuir-Blodgett multilayer films of side-chain liquid crystal copolymers

Abstract X-ray reflectivity studies on Langmuir-Blodgett multilayer films of side-chain liquid crystal polymers are reported. The films have a high degree of lamellar order. The layer periodicity is independent of the type of monolayer deposition, implying a reorientation of the side group mesogens following the deposition process. X-ray reflectivity from thin films displays subsidiary maxima permitting a quantitative measure of the change in side-chain density between multilayer and monolayer. A unit cell density profile is calculated for thick films assuming a symmetric unit cell.

Cloning of a rat homologue of mouse GlyCAM 1 reveals conservation of structural domains

Recently we described the isolation of a mouse cDNA clone encoding a mucin-like endothelial glycoprotein that appears to function as an adhesive ligand for L selectin. This ligand has been named GlyCAM 1 (Gly-cosylation-dependent Cell Adhesion Molecule 1) because its adhesive interactions with the L selectin lectin domain require that the GlyCAM 1 polypeptide chain be appropriately modified with carbohydrates. These carbohydrate modifications include the addition of sialic acid as well as sulfate residues to O-linked carbohydrate side chains that are clustered in two serine/threonine-rich domains of the mucin. An additional interesting structure that may have relevance to the association of GlyCAM 1 with the lumenal surface of the endothelium was a potential amphipathic helix at the C terminus of the glycoprotein. In order to examine the importance of the postulated O-linked domains as well as the potential amphipathic helix, we have cloned the rat homologue of GlyCAM 1. The sequence of this clone reveals a serine/threonine-rich protein that is highly homologous with the mouse GlyCAM 1. As was found for the mouse GlyCAM 1, the rat homologue shows a clustering of these potential O-linked carbohydrate acceptors in two domains of the protein. Interestingly, many of the serines and threonines are found to be spaced identically in the two homologues, consistent with the possibility that both density and position of the O-linked side chains may be important for appropriate L selectin-mediated adhesion. In support of its postulated functional importance, the C-terminal potential amphipathic helix is conserved in the rat homologue. Finally, immunoprecipitation analysis of [35S]sulfate-labeled rat lymph nodes with either a mouse L selectin IgG chimera or a peptide antiserum directed against a relatively conserved portion of mouse GlyCAM 1 demonstrates a approximately 45-kDa sulfated ligand in rat lymph nodes that is analogous to that previously described for mouse lymph nodes.

Synthesis and Properties of Charged Copolypeptides Membranes as Biodegradable Materials

Three-component random copolypeptides consisting of N-hydroxyalkyl L-glutamine, L-aspartic acid, and L-lysine, related two component random copolymers and homopolymers, were prepared by aminolysis with 2-amino-1-ethanol [AE] or 5-amino-1-pentanol [APe], followed by crosslinking reaction with 1,8-octamethylenediamine [OMDA] on starting polymer membranes consisting of γ-methyl L-glutamate [E], L-aspartic acid [D], and L-lysine [K]. All water soluble samples prepared without OMDA were degraded by random chain fracture with papain. The effective crosslink density was proportional to the content of the crosslinker [OMDA] in the reaction mixture. Tensile properties of these hydrophilic membranes were highly dependent on the swelling ratio in a pseudo-extracellular fluid [PECF], hydrophobicity of side chains. The effective charge density of membranes, and their behavior were typical of elastomers. A higher rate of water permeability was obtained with charged membranes than non-charged and/or compensated charged membranes with the same order of swelling ratio in PECF. Biodegradation of the samples in vitro by papain indicated that degradation could be regarded as a bulk rather than surface phenomenon. The rate of degradation was also dependent on the swelling ratio of membranes, as well as on the hydrophobicity and effective charge density of side chains of the samples.

Optically‐active comb‐branch liquid crystalline polymers having the chiral center in the flexible spacer unit. III. Copolymers of (S)‐2‐(4′‐cyano‐4‐biphenylyloxy)‐1‐methylethoxy ethyl methacrylate and di[6‐(4‐methoxy‐4′‐oxybiphenyl) hexyl]‐2‐methylene butane‐1,4‐dioate

AbstractA new series of copolymers with optically‐active liquid crystalline side chain units has been synthesized from the comonomers (S)‐2‐[2‐(4′‐cyano‐4‐biphenylyloxy‐1‐methylethoxy]ethyl methacrylate (1) and di[6‐(4‐methoxy‐4′‐oxybiphenyl)hexyl]‐2‐methylene butane‐1,4‐dioate (4). Chiral nematic phases were exhibited by two members of the series, rich in monomer 1, while a smectic phase was exhibited in copolymers rich in 4. While it was thought possible that ordered chiral liquid crystalline phases may be induced by copolymerizing chiral mesogenic monomers with mesogenic derivatives of itaconic acid where the high side chain density encourages greater ordering in the system, no evidence of smectic C* phases could be found in the present systems. © 1993 John Wiley & Sons, Inc.

The alignment of polar mesogens in electroactive polymers

We have studied the polar alignment produced in a range of comb polymers with a methacrylate backbone and oxynitrostilbene mesogenic groups attached to the backbone at variable densities by a 3 or 6 carbon spacer. These polymers, when poled, display second order non-linear optical activity, piezoelectricity and pyroelectricity. The magnitude of these effects depends upon the degree of polar ordering of the side groups and we have therefore investigated the efficiency of the poling process using the pyroelectric coefficient and thermally stimulated currents as primary diagnostic techniques. For constant poling conditions it is found that the polarization and pyroelectric response are not linearly related to the volume fraction of mesogen, i.e. samples of high mesogen concentration display lower polarization and pyroelectric activity than might be expected from extrapolation of low concentration data. Indeed, for the 6 carbon spacer material there would appear to be an optimum mesogen concentration around 30 mol%. Also, though the 3 and 6 carbon spacer materials are similar at low mesogen concentration, the latter material gives a lower response when it forms a liquid crystalline phase at high concentrations. We therefore suggest that the polar mesogens tend to pack anti-parallel at higher concentrations and that this interaction is not readily overcome by the applied field. Though these polymers are brittle at high mesogen concentration, ductile thin films can be cast if the density of side chains is reduced to 10 mol%. Despite the reduction in mesogen concentration, the pyroelectric activity is not proportionally reduced, implying more effective poling due to reduced interaction between the mesogens.

Crystal structure of cholesterol oxidase from Brevibacterium sterolicum refined at 1.8 Å resolution

Cholesterol oxidase (3β-hydroxysteroid oxidase, EC 1.1.3.6) is an FAD-dependent enzyme that carries out the oxidation and isomerization of steroids with a trans A: B ring junction. The crystal structure of the enzyme from Brevibacterium sterolicum has been determined using the method of isomorphous replacement and refined to 1.8 Å resolution. The refined model includes 492 amino acid residues, the FAD prosthetic group and 453 solvent molecules. The crystallographic R-factor is 15.3% for all reflections between 10.0 Å and 1.8 Å resolution. The structure is made up of two domains: an FAD-binding domain and a steroid-binding domain. The FAD-binding domain consists of three non-continuous segments of sequence, including both the N terminus and the C terminus, and is made up of a six-stranded β-sheet sandwiched between a four-stranded β-sheet and three α-helices. The overall topology of this domain is very similar to other FAD-binding proteins. The steroid-binding domain consists of two non-continuous segments of sequence and contains a six-stranded antiparallel β-sheet forming the “roof” of the active-site cavity. This large β-sheet structure and the connections between the strands are topologically similar to the substrate-binding domain of the FAD-binding protein para-hydroxybenzoate hydroxylase. The active site lies at the interface of the two domains, in a large cavity filled with a well-ordered lattice of 13 solvent molecules. The flavin ring system of FAD lies on the “floor” of the cavity with N-5 of the ring system exposed. The ring system is twisted from a planar conformation by an angle of approximately 17 °, allowing hydrogen-bond interactions between the protein and the pyrimidine ring of FAD. The amino acid residues that line the active site are predominantly hydrophobic along the side of the cavity nearest the benzene ring of the flavin ring system, and are more hydrophilic on the opposite side near the pyrimidine ring. The cavity is buried inside the protein molecule, but three hydrophobic loops at the surface of the molecule show relatively high temperature factors, suggesting a flexible region that may form a possible path by which the substrate could enter the cavity. The active-site cavity contains one charged residue, Glu361, for which the side-chain electron density suggests a high degree of mobility for the side-chain. This residue is appropriately positioned to act as the proton acceptor in the proposed mechanism for the isomerization step.

Soluble poly(para‐phenylene)s, 3. Variation of the length and the density of the solubilizing side chains

AbstractThe synthesis of a variety of alkyl‐chain‐substituted poly(para‐phenylene) (PPP) derivatives, including those with systematically decreased density of side chains, is described, using for their preparation the Pd‐catalyzed polymerization method recently developed in Mainz (Mainz route to PPP's). The degrees of polymerization achieved range between 30 and 50, allowing the synthesis of structurally well‐defined and soluble PPP's with more than hundred 1,4‐phenylene rings attached to each other.

Synthesis and properties of hydrophilic copolypeptide membranes containing L‐aspartic acid as one component

AbstractThree‐component random copolypeptide consisting of N‐hydroxypropyl‐L‐glutamine, L‐aspartic acid, and L‐lysine, related two component random copolypeptides and homopolypeptides, were prepared by carrying out aminolysis reaction with 3‐amino‐1‐propanol, followed by crosslinking reaction with 1,8‐octamethylenediamine on starting polymer membranes consisting of γ‐methyl‐L‐glutamate, L‐aspartic acid, and L‐lysine. The effective crosslink density was shown to be proportional to the content of the crosslinker in the reaction mixture. The tensile properties of these hydrophilic membranes were highly dependent on the degree of swelling in the pseudoextracellular fluid, hydrophobicity of the side chains, and the effective charge density of membranes, and their behavior was typical of an elastomer. A higher rate of water permeability was obtained with charged membranes than noncharged and/or compensated charged membranes with the same order of the degree of swelling in the pseudo‐extracellular fluid. Biodegradation of the samples in vitro by papain indicated that the degradation could be regarded as a bulk rather than a surface phenomenon. The rate of degradation was also highly dependent on the degree of swelling of membranes, as well as on the hydrophobicity and the effective charge density of side chains of sample membranes.

The improvement of Cα atom positions of trichosanthin—–An application of phase filtering procedure to three‐dimensional structure determination of protein

AbstractWang's phase filtering procedure has been applied to the structure study of trichosanthin crystals belonging to space group C2. The MIR phases at 3Å resolution have been improved with the accumulative phase shift of 31.8° and the figure of merit has been improved from 0.54 to 0.84. The resulted electron density map at 3Å resolution has been much improved with a higher ratio of the electron density of the protein to the noise in the solvent region. The minimap has been interpreted more reasonably than the previous maps, giving the overall course of the polypeptide chain substantially the same as previously reported except for containing more secondary structure and a few short segments traced differently. The first 85 Cα atoms of the polypeptide chain are located in about the same positions as determined previously with an exception of a short segment but the amino acid residues after that shift along the chain with a maximum shift of about 15 residues. The electron density of side chains agrees better with the primary structure. The conformation and the three‐dimensional structure as well as the major heavy‐atom binding sites for the two molecules in an asymmetric unit are substantially the same. The structure of trichosanthin shows more similarity to ricin A chain than that reported previously.

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.

Analysis of the carbohydrate-binding specificity of lectin-conjugated lipid vesicles which interact with polysaccharide fragments

In order to obtain some insight into understanding of the multivalent interactions between polysaccharide fragments as ligands and lectins conjugated on the membrane surface of lipid vesicles as receptors, two kinds of lectin-conjugated lipid vesicles were prepared, and studies on agglutination and binding were performed with two kinds of microscale assays by applying these to two model systems consisting of different ligands and receptors. The agglutination studies based on the interaction between plant galactomannan fragments and Ricinus communis agglutinin-conjugated lipid vesicles indicated three possible cases of interaction; strong agglutination was caused by large-size fragments with high side chain density, moderate agglutination by medium-size fragments with low side chain density and no agglutination by small-size fragments. The binding studies based on the interaction between yeast mannan fragments and concanavalin A lectin-conjugated lipid vesicles, together with glycosyl-linkage composition analyses, suggested that binding efficiency should be correlated with the structural characteristics of ligands such as size of fragments, rate of branching, and number of high-affinity residues.

Preparation and Properties of Charged Copolypeptide Membranes as Biodegradable Materials

Three component random copolypeptides consisting of N-hydroxyalkyl L-glutamine, L-glutamic acid, and L-lysine were prepared by carrying out aminolysis reactions with aminoalcohols, such as 3-amino-1-propanol (P) or 5-amino-1-pentanol (Pe), followed by crosslinking reaction with octamethylenediamine (OMDA) on starting polymer membranes consisting of γ-methyl-L-glutamate (M), L-glutamic acid (B), and L-lysine (K). The effective crosslink density was shown to be proportional to the content of the crosslinker (OMDA) in the reaction mixture. The tensile properties of these hydrophilic membranes were highly dependent on the degree of swelling in the pseudo-extracellular fluid (PECF), hydrophobicity of the side chains, and the effective charge density of membranes, and their behavior was typical of an elastomer. A higher rate of water permeability was obtained with charged membranes than non-charged and/or compensated charged membranes with the same order of the degree of swelling in PECF. Biodegradation of the samples in vitro by pronase E and papain indicated that the degradation could be regarded as a bulk rather than a surface phenomenon. The rate of degradation was also highly dependent on the degree of swelling of membranes, as well as on the hydrophobicity and effective charge density of side chains of the samples.

The structure of glycogen phosphorylase a at 2.5 Å resolution

The structure of the glucose-inhibited form of glycogen phosphorylase a has been determined at a resolution of 2.5 Å. With the aid of the primary sequence derived by Titani et al. (1977) for this enzyme, we have constructed an atomic model of the 97,400 molecular weight monomer. A substantial improvement in the electron density map over that reported previously (Fletterick et al., 1976 b) was achieved by extension of the data set to 2.5 Å and the inclusion of three additional “heavy-atom” derivatives in the phasing procedure. Main-chain and side-chain electron density are clearly resolved in the map, allowing an unambiguous correlation with the published primary structure. The course of the polypeptide backbone in the C-terminal half of the molecule has been modified at two positions from that reported in the 3.0 Å resolution interpretation. The enzyme is clearly organized into two domains, both with α β packing topology. The catalytic site lies in a crevice at the interface between the two domains. α- d-Glucose, which stabilizes the inactive (T) conformation in the parent crystal, is bound at this site in the C (6′) chair equatorial conformation within 6 Å of the pyridoxal phosphate coenzyme which is covalently bound through the ϵ-amino group of lysine 679. The larger, N-terminal domain is differentiated by folding architecture and tertiary contacts into two lobes or subdomains which share the same β-sheet backbone: the predominantly helical glycogen storage (maltoheptaose binding) lobe and the N-terminal subdomain. The latter is involved in a variety of protein-protein interactions with the monomer related by the 2-fold axis of the physiological dimer, and contains the serine 14-phosphate moiety and the AMP (positive effector) binding site. The core of the second domain is the complex (βαβ)′ folding unit previously characterized as the nucleotide binding fold (Rao & Rossmann, 1973).

Influence of side chain structure on sorption of xenon and carbon dioxide gas by polypeptide membranes

The influence of side chain structure on sorption of xenon and carbon dioxide gas by polypeptides is examined in two ways: chemical structure of side chain and higher order structure of the side chain region. Sorption of xenon gas increases progressively with increase in side chain length of poly(n-alkyl L-glutamates). The sorption isotherms of a polymer at various temperatures are reduced to one curve when the data are arranged in the form of sorption per residue versus fugacity ratio (as activity). The sorption of carbon dioxide gas shows a minimum at an intermediate length of side chains, corresponding to poly(n-propyl or n-butyl glutamate). The behavior is considered the result of overlapping influences of looseness of the side chain region and density of polar groups. The Langmuir constants and the enthalpy and entropy of sorption explain this quantitatively. Sorption of carbon dioxide gas is influenced by the higher-order structure of poly(γ-benzyl L-glutamate). In this polymer, the benzyl ester part is the main sorption site, and it was confirmed that the stacking between side-chain benzene rings significantly reduces the extent of sorption.

National pilot project

Coatings made with water dispersions of different nano-particles of photocatalytic additives (titania and titania doped with iron and vanadium) were prepared with diverse superplasticizers, SPs, to optimize the atmospheric NO removal efficiency when applied onto cement- and air-lime mortars. The use of different polycarboxylate-based superplasticizers (52IPEG, 23APEG and 45PC6) prevented nano-particles from agglomeration. The steric hindrance, provided by a large density and length of side chains, was ascertained as the most effective repulsion mechanism and 52IPEG was the most efficient SP. In PC- and air-lime mortars, the coatings with polycarboxylate-based SPs improved the NO removal rates as compared with the SP-free coating: an average increase of NO degradation by 15% under UV and by 76% under solar light was found. This finding was related to the drop in the agglomeration of the photocatalysts, with more exposed active sites and a decrease of the electron-hole recombination rates. Capillary water absorption and water vapour permeability values showed that the coatings did not alter the performance of the mortars. SEM examination showed that the use of SPs enhanced the distribution of the photocatalysts yielding thinner coating layers and boosting the percolation of the active material within the mortars. Accelerated weathering showed a moderate reduction of NO removal efficiency. Coating with 52IPEG was the most efficient in preserving the activity. Measurements of Ti content showed a low washout of the TiO2 nano-particles, supporting the long-run activity of these coatings.