Showing papers on "Circular dichroism published in 2002"

DICHROWEB: an interactive website for the analysis of protein secondary structure from circular dichroism spectra.

Abstract: A user-friendly website for the analysis of protein secondary structures from Circular Dichroism (CD) and Synchrotron Radiation Circular Dichroism (SRCD) spectra has been created.

The formation of polymer vesicles or "peptosomes" by polybutadiene-block-poly(L-glutamate)s in dilute aqueous solution.

Abstract: Polybutadiene-block-poly(l-glutamate) copolymers were made by anionic polymerization and subsequent ring-opening polymerization of N-carboxyanhydrides and were characterized by NMR, IR, SEC, and circular dichroism. These polymers, when appropriately designed, form so-called “polymersomes” or “peptosomes”, vesicles composed of modified protein units. The size and structure of the vesicles are determined by dynamic light scattering, small-angle neutron scattering, and freeze-fracture electron microscopy. It is also shown that the size of the peptosomes does not depend on the pH; that is, the solvating peptide units can perform a helix−coil transition without serious changes of the vesicle morphology.

Chiroptical properties from time-dependent density functional theory. I. Circular dichroism spectra of organic molecules

Abstract: We report the implementation of the computation of rotatory strengths, based on time-dependent density functional theory, within the Amsterdam Density Functional program. The code is applied to the simulation of circular dichroism spectra of small and moderately sized organic molecules, such as oxiranes, aziridines, cyclohexanone derivatives, and helicenes. Results agree favorably with experimental data, and with theoretical results for molecules that have been previously investigated by other authors. The efficient algorithms allow for the simulation of CD spectra of rather large molecules at a reasonable accuracy based on first-principles theory. The choice of the Kohn–Sham potential is a critical issue. It is found that standard gradient corrected functionals often yield the correct shape of the spectrum, but the computed excitation energies are systematically underestimated for the samples being studied. The recently developed exchange-correlation potentials “GRAC” and “SAOP” often yield much better a...

Is polyproline II a major backbone conformation in unfolded proteins

Abstract: Publisher Summary Protein folding is a process by which a polypeptide chain acquires its native structure from an unfolded state through a transition state. Recent studies of the unfolded states of proteins are based on a modification of the random coil model, recognizing that in many cases some residual native or non-native structure persists.. Combined evidence from the theoretical study of a blocked alanine peptide in aqueous solution and a variety of spectroscopic studies, including ultraviolet circular dichroism (CD), nuclear magnetic resonance (NMR), two-dimensional vibrational spectroscopy, vibrational circular dichroism (VCD), and vibrational Raman optical activity (VROA) reveal that the polyproline II (P II ) conformation is the dominant conformation in a variety of short model peptides. This chapter discusses the evidence from short peptides. It reviews the circular dichroism of unfolded proteins and addresses the role of P II in unfolded proteins.

Helical rosette nanotubes with tunable chiroptical properties.

Abstract: On the basis of transmission electron microscopy (TEM), dynamic light scattering (DLS), small-angle X-ray scattering (SAXS), and circular dichroism (CD) studies, compound 1 was shown to exist mainly in two states: (a) At high concentration (> or =1 mM, in methanol), 1 undergoes hierarchical self-assembly to generate rosette nanotubes with approximately 4 nm diameter and a concentration-dependent hydrodynamic radius in the range 10-100 nm. Under these conditions, addition of a chiral amino acid promoter (L-Ala), that binds to the crown ether moiety of 1 via electrostatic interactions, promotes a rapid transition (k(0) approximately equal 0.48 s(-1), for [1] = 0.046 mM, [L-Ala] = 2.8 mM) from racemic to chiral rosette nanotubes with predefined helicities as indicated by the resulting induced circular dichroism (ICD). (b) At low concentration (< or =0.04 mM, in methanol), 1 exists mainly in a nonassembled state as shown by TEM and DLS. Addition of L-Ala in this case triggers a relatively slow (k(0) approximately equal 0.07 s(-1) for [1] = 0.04 mM, [L-Ala] = 2.4 mM) sequence of supramolecular reactions leading to the hierarchical self-assembly of rosette nanotubes with predefined helicities. Under both conditions a and b, the kinetic data unveiled the intrinsic ability of the rosette nanotubes to promote their own formation (autocatalysis). The degree of chiral induction was found to depend dramatically upon the chemical structure of the promoter. This process appears also to follow an all-or-none response, as the vast majority of the crown ether sites must be occupied with a promoter for a complete transition to chiral nanotubes to take place. Finally, both supramolecular pathways a and b offer an efficient approach for the preparation of helical rosette nanotubes with tunable chiroptical properties and may also be viewed as a process by which a predefined set of physical and chemical properties that characterizes a molecular promoter is expressed at the macromolecular level.

Synthesis, characterization, and optical properties of monodisperse chiral oligofluorenes.

Abstract: The first series of monodisperse chiral oligofluorenes was synthesized and characterized. Chain length was found to play an important role in solid morphology. Whereas dimer through tetramer are amorphous, pentamer through hexadecamer all show cholesteric mesomorphism with varying degrees of morphological stability against crystallization. Pristine spin-cast films, approximately 90 nm in thickness, are amorphous but exhibit pronounced circular dichroism and highly efficient circularly polarized fluorescence, suggesting the presence of chiral assemblies that remain to be experimentally characterized. A nonamer with two sets of the 2S-methylbutyl group replaced by the 3S,7-dimethyloctyl group was prepared and shown to be capable of forming a monodomain, glassy cholesteric film with thermal treatment. The cholesteric film is responsible for an order-of-magnitude increase in circular dichroism and a handedness reversal in circularly polarized fluorescence as compared to the amorphous pristine film. Molecular dynamics simulation furnished new insight into the molecular origin of the observed chiral optical properties in neat films.

Spectroscopic Characterization of Nonnative Conformational States of Cytochrome c

Abstract: Protein and heme structural changes of ferric and ferrous cytochrome c (Cyt-c) that are induced by electrostatic binding (e.g., liposomes, electrodes), by hydrophobic interactions (e.g., monomeric sodium dodecyl sulfate), by guanidium hydrochloride (GuHCl), and at low pH and high temperature were studied by UV−vis absorption, circular dichroism (CD), electron paramagnetic resonance (EPR), and (surface-enhanced) resonance Raman [(SE)RR] spectroscopy. In a global spectral analysis, all species that differ with respect to the heme structure were identified and characterized in terms of the spin and ligation state of the heme as well as of protein secondary and tertiary structure changes. The results indicate that the upper part of the heme pocket including the Met-80 ligand is the most labile protein region such that this ligand is dissociated from the heme iron in all nonnative Cyt-c states. Among these states, there are two six-coordinated low-spin (LS) configurations with H2O or His-33 serving as the sixt...

Tripeptides adopt stable structures in water. A combined polarized visible Raman, FTIR, and VCD spectroscopy study.

Abstract: We have measured the band profile of amide I in the infrared, isotropic, and anisotropic Raman spectra of L-alanyl-D-alanyl-L-alanine, acetyl-L-alanyl-L-alanine, L-vanyl-L-vanyl-L-valine, L-seryl-L-seryl-L-serine, and L-lysyl-L-lysyl-L-lysine at acid, neutral, and alkaline pD. The respective intensity ratios of the two amide I bands depend on the excitonic coupling between the amide I modes of the peptide group. These intensity ratios were obtained from a self-consistent spectral decomposition and then were used to determine the dihedral angles between the two peptide groups by means of a recently developed algorithm (Schweitzer-Stenner, R. Biophys. J. 2002, 83, 523-532). The validity of the obtained structures were checked by measuring and analyzing the vibrational circular dichroism of the two amide I bands. Thus, we found two solutions for all protonation states of trialanine. Assuming a single conformer, one obtains a very extended beta-helix-like structure. Alternatively, the data can be explained by the coexistence of a 3(1)(PII) and a beta-sheet-like structure. Acetyl-L-alanyl-L-alanine exhibits a structure which is very similar to that obtained for trialanine. The tripeptide with the central D-alanine adopts an extended structure with a negative psi and a positive phi angle. Trivaline and triserine adopt single beta(2)-like structures such as that identified in the energy landscape of the alanine dipeptide. Trilysine appears different from the other investigated homopeptides in that it adopts a left-handed helix which at acid pD is in part stabilized by hydrogen bonding between the protonated carboxylate (donor) and the N-terminal peptide carbonyl. Our result provides compelling evidence for the capability of short peptides to adopt stable structures in an aqueous solution, which at least to some extent reflect the intrinsic structural propensity of the respective amino acids in proteins. Furthermore, this paper convincingly demonstrates that the combination of different vibrational spectroscopies provides a powerful tool for the determination of the secondary structure of peptides in solution.

Can one derive the conformational preference of a beta-peptide from its CD spectrum?

Abstract: CD spectroscopy is often used to elucidate the secondary structure of peptides built from non-natural amino acids such as beta-amino acids. The interpretation of such CD spectra is not always unambiguous. Here, we present a case where two beta-hexapeptides, a dimethyl-beta-hexapeptide indicated as DM-BHP (A) and its nonmethylated analogue indicated as BHP (B), exhibit similar CD spectra, whereas they are expected to differ in secondary structure. The structural properties of both peptides were studied by molecular dynamics simulation, and from the resulting trajectories, the corresponding CD spectra were calculated. Starting from a fully extended conformation, BHP is observed to form a 3(14)-helix, while DM-BHP remains unfolded. However, even though these two peptides hardly share any conformations, their calculated CD spectra are alike and show the same features as the experimentally measured ones. Our results imply that a particular CD pattern can be induced by spatially different structures, which makes it difficult to derive the conformational preference of a peptide from its CD spectrum alone. To gain more insight into the relationship between the preferred conformation of a peptide and its CD spectrum, more accurate methods to calculate the CD spectrum for a given conformation are required.

Size-dependent magnetism of deposited small iron clusters studied by x-ray magnetic circular dichroism.

Abstract: The size-dependent magnetic properties of small iron clusters deposited on ultrathin nickel films have been studied with circularly polarized synchrotron radiation. With the use of sum rules, orbital and spin magnetic moments have been extracted from x-ray magnetic circular dichroism spectra. The ratio of orbital to spin magnetic moments varies considerably with cluster size, reflecting the dependence of magnetic properties on cluster size and geometry. These variations can be explained in terms of enhanced orbital moments in small clusters.

Functional consequences of preorganized helical structure in the intrinsically disordered cell-cycle inhibitor p27(Kip1).

Abstract: p27Kip1 contributes to cell-cycle regulation by inhibiting cyclin-dependent kinase (Cdk) activity. The p27 Cdk-inhibition domain has an ordered conformation comprising an α-helix, a 310 helix, and β-structure when bound to cyclin A-Cdk2. In contrast, the unbound p27 Cdk-inhibition domain is intrinsically disordered (natively unfolded) as shown by circular dichroism spectroscopy, lack of chemical-shift dispersion, and negative heteronuclear nuclear Overhauser effects. The intrinsic disorder is not due to the excision of the Cdk-inhibition domain from p27, since circular dichroism spectra of the full-length protein are also indicative of a largely unfolded protein. Both the inhibition domain and full-length p27 are active as cyclin A-Cdk2 inhibitors. Using circular dichroism and proline mutagenesis, we demonstrate that the unbound p27 Cdk-inhibition domain is not completely unfolded. The domain contains marginally stable helical structure that presages the α-helix, but not the 310 helix, adopted upon bindin...

Mechanism of caveolin filament assembly.

Abstract: Caveolin-1 was the first protein identified that colocalizes with the ≈10-nm filaments found on the inside surface of caveolae membranes. We have used a combination of electron microscopy (EM), circular dichroism, and analytical ultracentrifugation to determine the structure of the oligomers that form when the first 101 aa of caveolin-1 (Cav1–101) are allowed to associate. We determined that amino acids 79–96 in this caveolin-1 fragment are arranged in an α-helix. Cav1–101 oligomers are ≈11 nm in diameter and contain seven molecules of Cav1–101. These subunits, in turn, are able to assemble into 50 nm long × 11 nm diameter filaments that closely match the morphology of the filaments in the caveolae filamentous coat. We propose that the heptameric subunit forms in part through lateral interactions between the α-helices of the seven Cav1–101 units. Caveolin-1, therefore, appears to be the structural molecule of the caveolae filamentous coat.

Contribution of Aromatic Interactions to α-Helix Stability

Abstract: The influence of natural and unnatural i, i + 4 aromatic side chain−side chain interactions on α-helix stability was determined in Ala-Lys host peptides by circular dichroism (CD). All interactions investigated provided some stability to the helix; however, phenylalanine−phenylalanine (F−F) and phenylalanine−pentafluorophenylalanine (F−f5F) interactions resulted in the greatest enhancement in helicity, doubling the helical content over i, i + 5 control peptides at internal positions. Quantification of these interactions using AGADIR multistate helix-coil algorithm revealed that the F−F and F−f5F interaction energies are equivalent at internal positions in the sequence (ΔGF-F = ΔGF-f5F = −0.27 kcal/mol), despite the differences in their expected geometries. As the strength of a face-to-face stacked phenyl−pentafluorophenyl interaction should surpass an edge-to-face or offset-stacked phenyl−phenyl interaction, we believe this result reflects the inability of the side chains in F−f5F to attain a fully stacke...

Chiroptical properties of chiral-substituted polyfluorenes

Abstract: Liquid-crystalline polyfluorene (PF) homopolymers substituted with chiral alkyl side chains were synthesized, and their chiroptical properties in the solid state were investigated by means of circular dichroism (CD), circularly polarized photoluminescence (CPPL), and circularly polarized electroluminescence (CPEL) measurements. Polarization-selective scattering of light is shown to cause artifacts in the circularly polarized absorption and emission spectra in the wavelength range near or above the absorption edge, and a measurement scheme to avoid these is presented. For all derivatives, significant chiroptical effects appeared only after the solid layers have been annealed at elevated temperatures, preferably into the liquid-crystalline state of the polymer. The largest anisotropy factors were measured for a polyfluorene substituted with chiral (R)-2-ethylhexyl side chains, yielding absolute values of up to 0.28 for CPPL and up to 0.25 for CPEL. These are among the highest ever reported for a chiral conj...

From chiral counterions to twisted membranes.

Abstract: In membranes, the chirality of the amphiphile constituents is sometimes expressed at a supramolecular scale of nanometers or micrometers. We have recently reported that membranes of nonchiral dicationic n−2−n amphiphiles can also be chirally twisted upon interacting with chiral tartrate counterions. Here, we demonstrate that the mechanism of the chiral induction by counterions involves specific anion−cation recognition and the induction of conformationally labile chirality in the cations. Single-crystal X-ray diffraction shows that the amphiphilic cations exist as a mixture of chiral conformers. 1H NMR data establish a specific recognition between tartrate and n−2−n cations and show that chiral conformers also exist in solution. Circular dichroism (CD) in the UV−vis shows a sharp conformational change of tartrate ions from anti to gauche when bound to the chiral cationic membranes. This is confirmed by CD in the infrared region which also shows concomitant induced CD bands in the vibrations of the n−2−n a...

Drug-membrane interactions : analysis, drug distribution, modeling

Abstract: Preface. Foreword. INTRODUCTION Function, Composition, and Organization of Membranes OCTANOL--WATER PARTITION VERSUS PARTITIONING INTO MEMBRANES ANALYTICAL TOOLS FOR THE ANALYSIS AND QUANTIFICATION OF DRUG--MEMBRANE INTERACTIONS High--performance Liquid Chromatography (HPLC) Displacement of 45Ca from Phospholipid Head Groups Differential Scanning Calorimetry (DSC) Fluorescence Techniques Fourier Transform Infrared Spectroscopy (FT--IR) Electron Spin Resoncance (ESR) Small Angle Neutron and X--ray Diffraction Nuclear Magnetic Resonance (NMR) Circular Dichroism (CD) UV Spectroscopy DRUG--MEMBRANE INTERACTIONS AND PHARMACOKINETICS OF DRUGS Drug Transport Drug Distribution Uptake into and Distribution within Bacterial Cells Drug Accumulation, Toxicity, and Selectivity DRUG--MEMBRANE INTERACTIONS AND PHARMACODYNAMICS Drug Efficacy Drug Resistance COMPUTER SIMULATION OF PHOSPHOLIPIDS AND DRUG--PHOSPHOLIPID INTERACTIONS Modeling Strategies for Studying Phospholipids and Drug--Phospholipid Interactions Computer Simulations with Phospholipids Concluding Remarks INDEX

Light-induced trimer to monomer transition in the main light-harvesting antenna complex of plants: thermo-optic mechanism.

Abstract: The main chlorophyll a/b light-harvesting complex of photosystem II, LHCIIb, has earlier been shown to be capable of undergoing light-induced reversible structural changes and chlorophyll a fluorescence quenching in a way resembling those observed in granal thylakoids when exposed to excess light [Barzda, V., et al. (1996) Biochemistry 35, 8981-8985]. The nature and mechanism of this unexpected structural flexibility has not been elucidated. In this work, by using density gradient centrifugation and nondenaturing green gel electrophoresis, as well as absorbance and circular dichroic spectroscopy, we show that light induces a significant degree of monomerization, which is in contrast with the preferentially trimeric organization of the isolated complexes in the dark. Monomerization is accompanied by a reversible release of Mg ions, most likely from the outer loop of the complexes. These data, as well as the built-in thermal and light instability of the trimeric organization, are explained in terms of a simple theoretical model of thermo-optic mechanism, effect of fast thermal transients (local T-jumps) due to dissipated photon energies in the vicinity of the cation binding sites, which lead to thermally assisted elementary structural transitions. Disruption of trimers to monomers by excess light is not confined to isolated trimers and lamellar aggregates of LHCII but occurs in photosystem II-enriched grana membranes, intact thylakoid membranes, and whole plants. As indicated by differences in the quenching capability of trimers and monomers, the appearance of monomers could facilitate the nonphotochemical quenching of the singlet excited state of chlorophyll a. The light-induced formation of monomers may also be important in regulated proteolytic degradation of the complexes. Structural changes driven by thermo-optic mechanisms may therefore provide plants with a novel mechanism for regulation of light harvesting in excess light.

γ2‐, γ3‐, and γ2,3,4‐Amino Acids, Coupling to γ‐Hexapeptides: CD Spectra, NMR Solution and X‐ray Crystal Structures of γ‐Peptides

Abstract: There are numerous possible γ-amino acids with different degrees of substitution and with various constitutions and configurations Of these the γ4- and the like- and unlike-γ2,4-amino acids have been previously used as building blocks in γ-peptides The synthesis of γ2-, γ3-, and γ2,3,4-peptides is now described The corresponding amino acids have been prepared by Michael addition of chiral N-acyl-oxazolidinone enolates to nitro-olefins, with subsequent reduction of the NO2 to NH2 groups Such additions to E-2-methyl-nitropropene provide (2R,3R,4R)-2-alkyl-3-methyl-4-amino-pentanoic acid derivatives (9, 10, 11) Stepwise coupling and fragment coupling lead to γ-di-, tri-, and hexapeptides (12–23), which were fully characterized The crystal structures of one of the γ-amino acids (2,3-dimethyl-4-amino-pentanoic acid⋅HCl, 9 a), of a γ2,3,4-di- and a γ2,3,4-tetrapeptide (20, 22) are described, and the NMR solution structure in MeOH of a γ2,3,4-hexapeptide (3) has been determined (using TOCSY, COSY, HSQC, HMBC and ROESY measurements and a molecular dynamics simulated-annealing protocol) A linear conformation (sheet-like), a novel (M) helix built of nine-membered hydrogen-bonded rings, and (M) 2614 helices have thus been identified NMR measurements at different temperatures (298–393 K) and H/D-exchange rates obtained for the γ2,3,4-hexapeptide are interpreted as evidence for the stability of the 2614 helix (no “melting”) and for its non-cooperative folding mechanism CD Spectra of the γ-peptides have been measured in MeOH and CH3CN, indicating that only the protected and unprotected γ2,3,4-hexapeptide is present as the 2614 helix in solution The structures of the γ2- and γ3-hexapeptides (1, 2) could not be determined

Temperature-Induced Extended Helix/Random Coil Transitions in a Group 1 Late Embryogenesis-Abundant Protein from Soybean

Abstract: Group 1 late embryogenesis-abundant (LEA) proteins are a subset of hydrophilins that are postulated to play important roles in protecting plant macromolecules from damage during freezing, desiccation, or osmotic stress. To better understand the putative functional roles of group 1 LEA proteins, we analyzed the structure of a group 1 LEA protein from soybean (Glycine max). Differential scanning calorimetry of the purified, recombinant protein demonstrated that the protein assumed a largely unstructured state in solution. In the presence of trifluoroethanol (50% [w/v]), the protein acquired a 30% α-helical content, indicating that the polypeptide is highly restricted to adopt α-helical structures. In the presence of sodium dodecyl sulfate (1% [w/v]), 8% of the polypeptide chain adopted an α-helical structure. However, incubation with phospholipids showed no effect on the protein structure. Ultraviolet absorption and circular dichroism spectroscopy revealed that the protein existed in equilibrium between two conformational states. Ultraviolet absorption spectroscopy studies also showed that the protein became more hydrated upon heating. Furthermore, circular dichroism spectral measurements indicated that a minimum of 14% of amino acid residues existed in a solvent-exposed, left-handed extended helical or poly (l-proline)-type (PII) conformation at 20°C with the remainder of the protein being unstructured. The content of PII-like structure increased as temperature was lowered. We hypothesize that by favoring the adoption of PII structure, instead of the formation of α-helical or β-sheet structures, group 1 LEA proteins retain a high content of surface area available for interaction with the solvent. This feature could constitute the basis of a potential role of LEA proteins in preventing freezing, desiccation, or osmotic stress damage.

Comparison of the Binding of Cadmium(II), Mercury(II), and Arsenic(III) to the de Novo Designed Peptides TRI L12C and TRI L16C

Abstract: Designed α-helical peptides of the TRI family with a general sequence Ac-G(LKALEEK)4G-CONH2 were used as model systems for the study of metal−protein interactions. Variants containing cysteine residues in positions 12 (TRI L12C) and 16 (TRI L16C) were used for the metal binding studies. Cd(II) binding was investigated, and the results were compared with previous and current work on Hg(II) and As(III) binding. The metal peptide assemblies were studied with the use of UV, CD, EXAFS, 113Cd NMR, and 111mCd perturbed angular correlation spectroscopy. The metalated peptide aggregates exhibited pH-dependent behavior. At high pH values, Cd(II) was bound to the three sulfurs of the three-stranded α-helical coiled coils. A mixture of two species was observed, including Cd(II) in a trigonal planar geometry. The complexes have UV bands at 231 nm (20 600 M-1 cm-1) for TRI L12C and 232 nm (22 600 M-1 cm-1) for TRI L16C, an average Cd−S bond length of 2.49 A for both cases, and a 113Cd NMR chemical shift at 619 ppm (CdI...

Circular dichroism spectra of short, fixed-nucleus alanine helices

Abstract: Very short alanine peptide helices can be studied in a fixed-nucleus, helix-forming system [Siedlicka, M., Goch, G., Ejchart, A., Sticht, H. & Bierzynski, A. (1999) Proc. Natl. Acad. Sci. USA 96, 903–908]. In a 12-residue sequence taken from an EF-hand protein, the four C-terminal peptide units become helical when the peptide binds La3+, and somewhat longer helices may be made by adding alanine residues at the C terminus. The helices studied here contain 4, 8, or 11 peptide units. Surprisingly, these short fixed-nucleus helices remain almost fully helical from 4 to 65°C, according to circular dichroism results reported here, and in agreement with titration calorimetry results reported recently. These peptides are used here to define the circular dichroism properties of short helices, which are needed for accurate measurement of helix propensities. Two striking properties are: (i) the temperature coefficient of mean peptide ellipticity depends strongly on helix length; and (ii) the intensity of the signal decreases much less rapidly with helix length, for very short helices, than supposed in the past. The circular dichroism spectra of the short helices are compared with new theoretical calculations, based on the experimentally determined direction of the NV1 transition moment.