Showing papers on "Circular dichroism published in 1992"
TL;DR: A new magneto-optical sum rule is derived for circular magnetic dichroism in the x-ray region (CMXD) and applications are discussed to transition-metal and rare-earth magnetic systems.
Abstract: A new magneto-optical sum rule is derived for circular magnetic dichroism in the x-ray region (CMXD). The integral of the CMXD signal over a given edge allows one to determine the ground-state expectation value of the orbital angular momentum. Applications are discussed to transition-metal and rare-earth magnetic systems.
2,041 citations
TL;DR: A new method based on neural network theory is presented to analyze and quantify the information content of far UV circular dichroism spectra, and it is demonstrated that for each wavelength interval a specific network is suitable.
Abstract: A new method based on neural network theory is presented to analyze and quantify the information content of far UV circular dichroism spectra. Using a backpropagation network model with a single hidden layer between input and output, it was possible to deduce five different secondary structure fractions (helix, parallel and antiparallel beta-sheet, beta-turn and random coil) with satisfactory correlations between calculated and measured secondary structure data. We demonstrate that for each wavelength interval a specific network is suitable. The remaining discrepancy between the secondary structure data from neural network prediction and crystallography may be attributed to errors in the determination of protein concentration and random noise in the CD signal, as indicated by simulations.
1,059 citations
TL;DR: Differences between the solution conformations of the beta-(1-39) and beta-1-42 peptides suggests that the last 3 C-terminal amino acids are crucial to amyloid deposition, and suggests that beta-peptide is not an intrinsically insoluble peptide.
664 citations
TL;DR: The structure of a synthetic peptide comprising the 28 amino-terminal residues of actin has been examined by 1H-NMR and CD spectroscopy and the significance of the TFE-induced peptide structure is discussed.
Abstract: The structure of a synthetic peptide comprising the 28 amino-terminal residues of actin has been examined by 1H-NMR and CD spectroscopy. The peptide is largely unstructured and flexible in solution but becomes increasingly structured at higher trifluoroethanol (TFE) concentrations. As judged by CD with the use of two additional peptides (actin 1-20 and actin 18-28), TFE induces formation of up to 48% helical content within residues 1-20, while residues 21-28 exhibit no helical propensity. Similar results were obtained by using NMR-derived distance information in restrained molecular dynamics calculations. The calculated structure of actin 1-28 peptide in 80% TFE is well defined for the first 23 residues with a backbone root mean square deviation of 0.5 A. Two helices are formed from residues 4-13 and 16-20, and a beta-turn is formed from residues 13-16. The N-terminal residues 1-3 exhibit increased flexibility and a helix-like conformation while the C-terminal residues 21-28 show no regular secondary structure. These results are compared with the predicted secondary structure and the structure of the corresponding sequence in the crystal structure of actin [Kabsch et al. (1990) Nature 347, 37-44]. The significance of the TFE-induced peptide structure is discussed.
573 citations
TL;DR: The results suggest that both oligomerization and ordered structure are necessary for biological activity and provide insights into the role of this region in HIV infection and the potential for development of a new class of antiviral agents.
Abstract: A peptide designated DP-107 was synthesized containing amino acid residues 558-595 of the envelope glycoprotein gp160 of human immunodeficiency virus type 1 strain LAI (HIV-1LAI). Algorithms for secondary structure have predicted that this region of the envelope transmembrane protein should form an extended alpha-helix. Consistent with this prediction, analysis by circular dichroism (CD) indicated that, under physiological conditions, DP-107 is approximately 85% helical. The high degree of stable secondary structure in a synthetic peptide of this size suggests self-association typical of a coiled coil or leucine zipper. In biological assays, the peptide efficiently blocked virus-mediated cell-cell fusion processes as well as infection of peripheral blood mononuclear cells by both prototypic and primary isolates of HIV-1. A single amino acid substitution in the peptide greatly destabilized its solution structure as measured by CD and abrogated its antiviral activity. An analogue containing a terminal cysteine was oxidized to form a dimer, and this modification lowered the dose required for antiviral effect from 5 to about 1 microgram/ml. These results suggest that both oligomerization and ordered structure are necessary for biological activity. They provide insights also into the role of this region in HIV infection and the potential for development of a new class of antiviral agents.
562 citations
TL;DR: The structural rearrangements in the protein molecules upon adsorption and subsequent displacement by morpholine were probed by determining the α-helix content from the circular dichroism of the proteins as discussed by the authors.
384 citations
TL;DR: Intermolecular association of short telomeric sequences reported here provides a possible model for chromosomal pairing and the formation of hairpin G quartet structure for the above sequences is supported by the enhanced electrophoretic mobility observed on non-denaturing polyacrylamide gels.
Abstract: The role of thymine residues in the formation of G-quartet structures for telomeric sequences has been investigated using model oligonucleotides of the type d(G4TnG4), with n = 1-4. Sequences d(G4T3G4) and d(G4T4G4) adopt a G-quartet structure formed by hairpin dimerization in 70 mM NaCl as judged by a characteristic circular dichroism signature with a 295 nm positive and 265 nm negative bands while d(G4TG4) adopts a parallel G-quartet structure like d(G12) which exhibits a strong positive band at 260 nm and a negative band at 240 nm. The sequence d(G4T2G4) exhibits a mixture of both conformations. The stability of hairpin G-quartet structures decreases with decrease in the number of intervening thymine residues. Potassium permanganate, a single strand specific probe has been used to establish the presence of loops composed of T residues in the hairpin G quartet structures formed by the oligonucleotides d(G4TnG4) with n = 2-4 in 70 mM NaCl. The formation of hairpin G quartet structure for the above sequences is further supported by the enhanced electrophoretic mobility observed on non-denaturing polyacrylamide gels. Human telomeric sequence d(TTAGGG)4 which showed enhanced electrophoretic mobility like Tetrahymena telomeric sequence d(T2G4)4 also exhibited a characteristic CD spectrum for a folded-back G-quartet structure. A detailed model for G-quartet structure involving hairpin dimer with alternating syn-anti-syn-anti conformation for the guanine residues both along the chain as well as around the G tetrad with at least two thymine residues in the loop is proposed. Intermolecular association of short telomeric sequences reported here provides a possible model for chromosomal pairing.
377 citations
TL;DR: Analysis of the difference between the ellipticity in benign buffer and in 50% trifluoroethanol and the slope term from a plot of the free energy of unfolding versus guanidine hydrochloride concentration supported the conclusion that the leucine residues at the ends of the coiled-coil are much less buried than in the middle section of thecoil.
308 citations
TL;DR: The chapter provides examples from laboratory of CD spectral changes in the near ultraviolet range that illustrate the CD characteristics of structural transitions in natural and synthetic DNAs.
Abstract: Publisher Summary Circular dichroism (CD) measurements are used to study the conformations of nucleic acids in solution. The reliance on CD spectroscopy to study DNA conformations has stemmed from the sensitivity and ease of CD measurements, the nondestructive nature of such measurements, the fact that conformations can be studied in solution, and the requirement for relatively small amounts of material. Although detailed structural information, such as from X-ray crystallography or NMR spectroscopy, is not available from CD spectra, the CD spectrum of DNA in solution can provide a reliable determination of its overall conformational state when compared with the CD spectra of reference samples. Moreover, CD spectroscopy is applicable to a wide range of samples, including those that are difficult to crystallize or to obtain at high concentrations. The chief practical application of CD spectroscopy to the study of DNA structures has been by making empirical comparisons with the CD spectra of known structures. The chapter provides examples from laboratory of CD spectral changes in the near ultraviolet range that illustrate the CD characteristics of structural transitions in natural and synthetic DNAs.
217 citations
TL;DR: The fluorescence-detected stopped-flow folding kinetics of ferricytochrome c exhibits a 35% decrease in fluorescence during the dead time, suggesting that a substantial decrease in the average tryptophan-heme distance occurs on a submillisecond time scale.
Abstract: The kinetics of protein folding for horse ferricytochrome c was investigated by stopped-flow methods, using far-UV circular dichroism (CD), near-UV CD, and tryptophan fluorescence to probe the formation of secondary structure and tertiary interactions. In the far-UV region of the CD spectrum (222 nm), 44% of the total change associated with refolding occurs within the dead time of the stopped-flow experiment, indicating that a significant amount of helical secondary structure is formed in less than 4 ms. The remaining changes in the ellipticity at 222 nm occur in two kinetic phases with time constants of about 40 ms and 0.7 s, respectively. In contrast, there is no evidence for rapid changes in the ellipticity at 289 nm: an aromatic CD band, which is indicative of the formation of a tightly packed core, only begins to appear in a 400-ms step and is completed in a final 10-s phase. The fluorescence of a single tryptophan at position 59, which becomes quenched upon folding via nonradiative energy transfer to the heme group, provides complementary information on the condensation of the polypeptide chain during refolding. The fluorescence-detected stopped-flow folding kinetics of ferricytochrome c exhibits a 35% decrease in fluorescence during the dead time, suggesting that a substantial decrease in the average tryptophan-heme distance occurs on a submillisecond time scale. The subsequent fluorescence changes exhibit two prominent phases with time constants of about 20 and 300 ms, followed by a minor 5-s phase. Transient peptide CD spectra measured at different folding times (4 ms to 5 s) show no evidence for non-native elements of secondary structure at any stage of folding. Together with previous pulsed amide proton exchange data measured under identical folding conditions (Roder, Elove, & Englander (1988) Nature 335, 700-7041, the results suggest that during the early stages of folding (<4 ms), a partially condensed intermediate with a fluctuating core is formed that contains a significant amount of helical secondary structure but no stable hydrogen bonds. A second intermediate, populated on the 20-1 00-ms time scale, exhibits stable hydrogen-bonded structure in two helical segments near the chain termini and increased compactness, but the aromatic side chains are still in a dynamic or exposed environment. Other helical segments contribute to the helical character of the far-UV CD spectrum but lack sufficiently stable hydrogen bonding to provide protection against NH exchange.
213 citations
TL;DR: The experiments suggest that the more likely peptide geometry is a 310-helix, and single and doubly spin-labelled analogues of alanine-based peptides in which the nitroxide spin label forms an unbranched side chain extending from the sulphur atom of a cysteine residue are designed.
Abstract: SHORT alanine peptides, containing 16 or 17 residues, appear to form α-helices in aqueous solution1–4. But the main spectroscopic analyses used on helical peptides (circular dichroism5 and nuclear magnetic resonance6–8) cannot distinguish between an α-helix (in which the ith residue is hydrogen-bonded to residue i+4; ref. 9) and the next most common peptide helix, the 310-helix10 (i → i + 3 hydrogen-bonding). To address this problem we have designed single and doubly spin-labelled analogues of alanine-based peptides in which the nitroxide spin label forms an unbranched side chain extending from the sulphur atom of a cysteine residue. Here we report the circular dichroism, Fourier-transform infrared and electron-spin resonance spectra of these peptides under helix-forming conditions. The infrared absorbance gives an amide I' band with a frequency that is substantially different from that observed for α-helices. The electron-spin resonance spectra of doubly labelled helices show that the ranking of distances between side chains, around a single turn (residues 4–8), is inconsistent with an α-helical structure. Our experiments suggest that the more likely peptide geometry is a 310-helix.
TL;DR: These findings suggest that it may act to unfold regions of ribosomal RNA so that a second domain of nucleolin has access to its specific binding site.
TL;DR: A 91 amino acid fragment of the heat shock transcription factor from both Saccharomyces cerevisiae and Kluyveromyces lactis contains a domain which forms a trimer via a triple-stranded alpha-helical coiled-coil, similar to that found in influenza hemagglutinin.
Abstract: We have isolated and characterized a 91 amino acid fragment of the heat shock transcription factor from both Saccharomyces cerevisiae and Kluyveromyces lactis. The two protein fragments behave similarly: they form homotrimers, as indicated by sedimentation equilibrium and cross-linking, and contain approximately 80% alpha-helix, as indicated by circular dichroism. Sedimentation velocity and diffusion coefficients indicate that they have an elongated, nonspherical shape. We conclude the following: these fragments contain a domain which forms a trimer via a triple-stranded alpha-helical coiled-coil, similar to that found in influenza hemagglutinin.
TL;DR: In this paper, the truncated staphylococcal nuclease was characterized by nuclear magnetic resonance, circular dichroism, and small-angle x-ray scattering measurements, and it was shown that extensive solvent exclusion generates a compact polypeptide chain prior to the development of persistent secondary structural features as a protein folds during biosynthesis.
Abstract: Deletion of 13 amino acids from the carboxyl terminus of the 149-amino acid staphylococcal nuclease molecule results in a denatured, partly unfolded molecule that lacks persistent secondary structure but is compact under physiological conditions. Since the modification is a carboxyl-terminal deletion, it is argued that the state resembles a peptide emerging from the ribosome just before the complete folding pathway is initiated. In this paper, we characterize the molecule by nuclear magnetic resonance, circular dichroism, and small-angle x-ray scattering measurements. The truncated nuclease shows wild-type levels of activity in the presence of calcium and is found to fold into a native-like conformation in the presence of 3',5'-bisphospho-2'-deoxythymidine, a potent inhibitor. Thus, the truncated molecule retains the capacity to fold. Our results suggest that extensive solvent exclusion generates a compact polypeptide chain prior to the development of persistent secondary structural features as a protein folds during biosynthesis.
TL;DR: It is concluded that the N terminus of HIV-1 IN contains a metal-binding domain whose structure is stabilized by tetrahedral coordination of metal by histidines 12 and 16 and cysteines 40 and 43.
TL;DR: The sequence homology within apolipoproteins A-I, E-3, and A-IV is used to derive a consensus sequence for two 11 amino acid repeating sequences in this family of proteins.
TL;DR: In this article, the major chlorophyll a/b light harvesting complex LHC-II was isolated from spinach thylakoid membranes after solubilization with the non-ionic detergent n-dodecyl-β- d -maltoside.
TL;DR: A mathematical model based on diffusion-limited aggregation was developed to describe the kinetics of aggregation and showed that β(1–40) aggregated end-to-end in solution, reaching average molecular weights greater than 4 × 106 after 13 days.
TL;DR: Ligand binding studies indicated some affinity of N9‐N2 for the triple helical region of collagen VI suggesting a role of the N‐terminal globule in the self‐assembly of microfibrils, but no or only little binding was observed to fibril‐forming collagens I and III, several basement membrane proteins and other extracellular proteins.
Abstract: A large portion of the N-terminal globule of human collagen VI was prepared from the culture medium of stably transfected human embryonic kidney cell clones. The recombinant product corresponds to sequence positions 1-1586 of the alpha 3 (VI) chain that consists of eight homologous approximately 200 residue motifs (N9 to N2) being similar to the A domain motif of von Willebrand factor. By ultracentrifugation fragment N9-N2 showed a molecular mass of 180 kDa and an asymmetric shape. Elongated structures that consist of eight small globes (diameter approximately 5 nm) were demonstrated by electron microscopy. The data indicate that each A domain motif represents a separate folding unit which are connected to each other by short protease-sensitive peptide segments. Circular dichroism studies demonstrated about 38% alpha helix, 14% beta sheets and 17% beta turns. Fragment N9-N2 showed binding to heparin which could be abolished by moderate salt concentrations. Heparin binding was assigned to domains N9, N6 and N3 which were obtained after partial proteolysis. Domains N7, N5 and N4 lacked affinity for heparin. In addition, N9-N2 showed strong binding to hyaluronan that required exposure to 6 M urea for full dissociation. Ligand binding studies indicated some affinity of N9-N2 for the triple helical region of collagen VI suggesting a role of the N-terminal globule in the self-assembly of microfibrils. No or only little binding was, however, observed to fibril-forming collagens I and III, several basement membrane proteins and other extracellular proteins. Fragment N9-N2 was also an inactive substrate for cell adhesion.
TL;DR: In this article, the authors examined the thermal stability of horse cytschrome c at low pH between 2.0 and 3.2 and different potassium chloride concentrations by absorbance of the Soret band, far and near-ultraviolet circular dichroism (u.v.c.
TL;DR: The kinetics of regain of the native ellipticity in the far- and near-UV spectra have been investigated and the spectrum was found to closely fit that predicted from the secondary structure of native lysozyme, suggesting that the molecular events observed in the two regions of the UV spectrum are either identical or strongly coupled.
Abstract: The kinetics of regain of the native ellipticity in the far- and near-UV spectra have been investigated during the refolding at pH 7.8 and 20 degrees C of guanidine-unfolded, nonreduced hen egg white lysozyme. Stopped-flow studies showed that the ellipticities at 260 and 289.5 nm exhibit biphasic kinetics with rate constants of about 50 s-1 and 2.5 s-1 for the rapid and slow phase, respectively. The ellipticity in the far-UV obeyed triphasic kinetics. In addition to a rapid and a slow phase with rate constants similar to those observed in the near-UV, a "burst" of ellipticity was shown to occur in the dead time of the experiments. The effects of low pH and of concentrations of guanidine ranging from 0.075 to 1.5 M on the rapid and slow rate constants were studied. Under all conditions investigated, the rate constants observed in the far- and near-UV for a given phase were the same, thus suggesting that the molecular events observed in the two regions of the UV spectrum are either identical or strongly coupled. Continuous-flow experiments at different wavelengths between 214 and 240 nm under conditions where the dead time for the observation was only 4 ms, followed by a detailed analysis of the kinetics of ellipticity change at each wavelength, provided the spectrum of the molecular species formed at the end of the burst phase. This spectrum was found to closely fit that predicted from the secondary structure of native lysozyme.(ABSTRACT TRUNCATED AT 250 WORDS)
TL;DR: Recombinant human myoglobin mutants with the distal His residue replaced by Leu, Val, or Gln residues were prepared by site-directed mutagenesis and expression in Escherichia coli to study the ligand binding properties.
TL;DR: The potential intervention of psRR‐DNA in recombination, gene expression and the stabilization of genomic structure is discussed, and a model derived from force‐field analysis in which the parallel‐stranded d(G‐A)n helix is right‐handed and constituted of alternating, symmetrical Gsyn and Aanti is proposed.
Abstract: The oligonucleotides d[(G-A)7G] and d[(G-A)12G] self-associate under physiological conditions (10 mM MgCl2, neutral pH) into a stable double-helical structure (psRR-DNA) in which the two polypurine strands are in a parallel orientation in contrast to the antiparallel disposition of conventional B-DNA. We have characterized psRR-DNA by gel electrophoresis, UV absorption, vacuum UV circular dichroism, monomer-excimer fluorescence of oligonucleotides end-labelled with pyrene, and chemical probing with diethyl pyrocarbonate and dimethyl sulfate. The duplex is stable at pH 4-9, suggesting that the structure is compatible with, but does not require, protonation of the A residues. The data support a model derived from force-field analysis in which the parallel-stranded d(G-A)n helix is right-handed and constituted of alternating, symmetrical Gsyn.Gsyn and Aanti.Aanti base pairs with N1H...O6 and N6H...N7 hydrogen bonds, respectively. This dinucleotide structure may be the source of a negative peak observed at 190 nm in the vacuum UV CD spectrum, a feature previously reported only for left-handed Z-DNA. The related sequence d[(GAAGGA)4G] also forms a parallel-stranded duplex but one that is less stable and probably involves a slightly different secondary structure. We discuss the potential intervention of psRR-DNA in recombination, gene expression and the stabilization of genomic structure.
TL;DR: The CD spectrum changes upon adsorption were significant in the “soft” protein molecules (myoglobin, hemoglobin, and BSA), while they were insingnificant in the“rigid” proteins (RNase A and peroxidase).
Abstract: The conformational changes in well-characterized model proteins [bovine ribonuclease A (RNase A), horseradish peroxidase, sperm-whole myoglobin, human hemoglobin, and bovine serum albumin (BSA)] upon adsorption on ultrafine polystyrene (PS) particles have been studied using circular dichroism (CD) spectroscopy. These proteins were chosen with special attention to molecular flexibility. The ultrafine PS particles were negatively charged and have average diameters of 20 or 30 nm. Utilization of these ultrafine PS particles makes it possible to apply the CD technique to determine the secondary structure of proteins adsorbed on the PS surface. Effects of protein properties and adsorption conditions on the extent of the changes in the secondary structure of protein molecules upon adsorption on ultrafine PS particles were studied. The CD spectrum changes upon adsorption were significant in the "soft" protein molecules (myoglobin, hemoglobin, and BSA), while they were insignificant in the "rigid" proteins (RNase A and peroxidase). The soft proteins sustained a marked decrease in alpha-helix content upon adsorption. Moreover, the native alpha-helix content, which is given as the percentage of the alpha-helix content in the free proteins, of adsorbed BSA was found to decrease with decreasing pH and increase with increasing adsorbed amount. These observations confirm some well-known hypotheses for the confirmational charges in protein molecules upon adsorption.
TL;DR: Resorcinol-dodecanal cyclotetramer 1 as an achiral host in chloroform forms hydrogen-bonded complexes with a variety of chiral di(poly)ols (2-24) including steroids and sugars as discussed by the authors.
Abstract: Resorcinol-dodecanal cyclotetramer 1 as an achiral host in chloroform forms hydrogen-bonded complexes with a variety of chiral di(poly)ols (2-24) including steroids and sugars. The complexation processes can be followed very conveniently by the induced circular dichroism (CD) spectroscopy. The binding constants as determined by CD titration increase in the order 15 (steroidal monool, K=8.7 M -1 )<3, 6, and 8 (acyclic glycols, (4.9-7.1×10)<9a, 10α, 12, and 13 (cyclic glycols and steroidal diols, (0.94-2.7)×10 2 )<11 (STEROIDAL TRIOL, 6.9×10 2 )
TL;DR: It is shown that the apo form of human retinol-binding protein (RBP) at neutral pH is in a rigid state with properties similar to those of holo-RBP, and that, at equilibrium, the pH-inducedretinol release from holo -RBP parallels denaturation of the apoprotein.
Abstract: Using far- and near-UV circular dichroism, viscosity, tryptophan fluorescence, NMR spectra, binding of a hydrophobic probe, and microcalorimetry, we have shown that the apo form of human retinol-binding protein (RBP) at neutral pH is in a rigid state with properties similar to those of holo-RBP. On the contrary, at acidic pH apo-RBP is in the molten globule state which has been earlier revealed for a number of proteins under mild denaturing conditions. We have also shown that, at equilibrium, the pH-induced retinol release from holo-RBP parallels denaturation of the apoprotein. These findings are consistent with our hypothesis that the transformation of RBP into the molten globule state is involved in the mechanism whereby retinol is delivered to target cells. In particular, a local acidic pH near the membrane surface of target cells might cause the transition of RBP to the molten globule state as well as the release of retinol.
TL;DR: Results indicate that DS500 (500-kDa dextran sulfate) induces aggregation of factor XII, and implies that aggregation provides a special mechanism whereby proteolytic cleavage is accomplished efficiently when factor XII molecules are bound side by side on the DS500 molecule.
TL;DR: Static fluorescence quenching measurements indicate that the helical axis of a representative lipid-associated peptide is, on average, oriented parallel to the surface of the membrane and located a few angstroms below the polar head group/hydrocarbon boundary.
Abstract: A 21-residue peptide of the sequence (LSSLLSL)3 forms ion channels when incorporated into planar lipid bilayer membranes of diphytanoylphosphatidylcholine (diPhy-PC). The frequency of channel openings increases with the applied voltage gradient. We investigated the molecular and structural mechanisms underlying this voltage dependence. A series of seven peptides, each containing a tryptophan substituted for a single residue in the middle heptad, was synthesized, purified, and incorporated into small, unilamellar, diPhy-PC vesicles. We measured circular dichroism, maximum fluorescence emission wave-lengths, and fluorescence quenching by both aqueous and lipid hydrocarbon-associated quenchers. Circular dichroism spectra and the observed sequence periodicity of all fluorescence and fluorescence quenching data are consistent with an alpha-helical peptide secondary structure. Energy transfer quenching measurements using N-terminally labeled (LSSLLSL)3 co-incorporated at lipid/peptide ratios greater than 100 into vesicles with one of the Trp-substituted peptides showed that the vesicle-associated peptide, in the absence of a voltage gradient across the bilayer, exists as an equilibrium mixture of monomers and dimers. Static fluorescence quenching measurements using different lipid-bound quenchers indicate that the helical axis of a representative lipid-associated peptide is, on average, oriented parallel to the surface of the membrane and located a few angstroms below the polar head group/hydrocarbon boundary. This surface orientation for the peptide is confirmed by the complementary sequence periodicity observed for Trp fluorescence emission wavelength shifts and collisional quenching by aqueous CsCl.(ABSTRACT TRUNCATED AT 250 WORDS)
TL;DR: Analysis of the secondary structure as a function of pH in two designed synthetic peptides with identical composition but different sequences, affords a quantitative estimate of the free energy contribution of a single ion pair to the stability of an isolated alpha-helix.