Showing papers on "Circular dichroism published in 2001"
TL;DR: Conjugates of bovine serum albumin and CdTe nanoparticles capped with l-cysteine have been synthesized via a one-pot glutaric dialdehyde cross-linking procedure and circular dichroism spectroscopy demonstrates that the tertiary structure of the protein remains largely intact after the conjugation.
Abstract: Conjugates of bovine serum albumin and CdTe nanoparticles capped with l-cysteine have been synthesized via a one-pot glutaric dialdehyde cross-linking procedure. Diads (1:1) with some amount of 2:1 albumin−nanoparticle assemblies preferably form in this reaction, as evidenced by gel electrophoresis. Circular dichroism spectroscopy demonstrates that the tertiary structure of the protein remains largely intact after the conjugation. Attachment of protein moieties result in a significant increase of CdTe emission, which is attributed to the resonance energy transfer from the tryptophan moieties of albumin to CdTe nanoparticles acting as receptors for the protein antennae.
410 citations
TL;DR: Silica entrapment is demonstrated to be fully compatible with structure analysis by circular dichroism, allows conformational studies in contact with solvents that would otherwise promote aggregation in solution, and generally enhances thermal protein stability.
Abstract: The sol-gel method of encapsulating proteins in a silica matrix was investigated as a potential experimental system for testing the effects of molecular confinement on the structure and stability of proteins. We demonstrate that silica entrapment (1) is fully compatible with structure analysis by circular dichroism, (2) allows conformational studies in contact with solvents that would otherwise promote aggregation in solution, and (3) generally enhances thermal protein stability. Lysozyme, α-lactalbumin, and metmyoglobin retained native-like solution structures following sol-gel encapsulation, but apomyoglobin was found to be largely unfolded within the silica matrix under control buffer conditions. The secondary structure of encapsulated apomyoglobin was unaltered by changes in pH and ionic strength of KCl. Intriguingly, the addition of other neutral salts resulted in an increase in the α-helical content of encapsulated apomyoglobin in accordance with the Hofmeister ion series. We hypothesize that protein conformation is influenced directly by the properties of confined water in the pores of the silica. Further work is needed to differentiate the steric effects of the silica matrix from the solvent effects of confined water on protein structure and to determine the extent to which this experimental system mimics the effects of crowding and confinement on the function of macromolecules in vivo.
395 citations
TL;DR: In this paper, a series of polyelectrolyte−surfactant complexes, DNA−cationic surfactant complexes and their self-assembled bulk film materials were prepared on a large scale.
Abstract: Series of polyelectrolyte−surfactant complexes, DNA−cationic surfactant complexes (cetyltrimethylammonium, cetylpyridinium, and cetylbenzyldimethylammonium), and their self-assembled bulk film materials were prepared on a large scale. Circular dichroism (CD) analysis indicated that the right-handed double helix structure of DNA was retained in these bulk film materials. TGA analysis suggested that 4 molecules of water were required to retain the B-type conformation of the DNA helix in the self-assembled bulk film materials. In addition, it revealed that DNA and the DNA−surfactant complex film materials were thermostable up to as high as 180 °C. Thermodynamical analysis indicated that these film materials were thermo-extensive over a temperature range from 100 to 148 °C. The DNA conformation in the supramolecular complex films can be reversibly tuned by changing the environmental humidity. Film formation was found to occur by self-assembly and self-organization with evaporation of solvent molecules. Variou...
357 citations
TL;DR: The marked increase in the lag time for fibril formation with mutations to more polar residues, as well as mutations to charged residues, demonstrates the importance of both hydrophobic and electrostatic interactions in the initial stages of fibrillation.
Abstract: The molecular basis of insulin fibril formation was investigated by studying the structural properties and kinetics of fibril formation of 20 different human insulin mutants at both low pH (conditions favoring monomer/dimer) and at pH 7.4 (conditions favoring tetramer/hexamer). Small-angle X-ray scattering showed insulin to be monomeric in 20% acetic acid, 0.1 M NaCl, pH 2. The secondary structure of the mutants was assessed using far-UV circular dichroism, and the tertiary structure was determined using near-UV circular dichroism, quenching of intrinsic fluorescence by acrylamide and interactions with the hydrophobic probe 1-anilino-8-naphthalene-sulfonic acid (ANS). The kinetics of fibril formation were monitored with the fluorescent dye, Thioflavin T. The results indicate that the monomer is the state from which fibrils arise, thus under some conditions dissociation of hexamers may be rate limiting or partially rate limiting. The insulin mutants were found to retain substantial nativelike secondary and tertiary structure under all conditions studied. The results suggest that fibril formation of the insulin mutants is controlled by specific molecular interactions that are sensitive to variations in the primary structure. The observed effects of several mutations on the rate of fibril formation are inconsistent with a previously suggested model for fibrillation [Brange, J., Whittingham, J., Edwards, D., Youshang, Z., Wollmer, A., Brandenburg, D., Dodson, G., and Finch, J. (1997) Curr. Sci. 72, 470-476]. Two surfaces on the insulin monomer are identified as potential interacting sites in insulin fibrils, one consisting of the residues B10, B16, and B17 and the other consisting of at least the residues A8 and B25. The marked increase in the lag time for fibril formation with mutations to more polar residues, as well as mutations to charged residues, demonstrates the importance of both hydrophobic and electrostatic interactions in the initial stages of fibrillation. A model for insulin fibril formation is proposed in which the formation of a partially folded intermediate is the precursor for associated species on the pathway to fibril formation.
327 citations
TL;DR: The data support the hypothesis that amyloid fibril formation involves the ordered self-assembly of partially folded species that are critical soluble precursors of fibrils.
Abstract: Light chain, or AL, amyloidosis is a pathological condition arising from systemic extracellular deposition of monoclonal immunoglobulin light chain variable domains in the form of insoluble amyloid fibrils, especially in the kidneys. Substantial evidence suggests that amyloid fibril formation from native proteins occurs via a conformational change leading to a partially folded intermediate conformation, whose subsequent association is a key step in fibrillation. In the present investigation, we have examined the properties of a recombinant amyloidogenic light chain variable domain, SMA, to determine whether partially folded intermediates can be detected and correlated with aggregation. The results from spectroscopic and hydrodynamic measurements, including far- and near-UV circular dichroism, FTIR, NMR, and intrinsic tryptophan fluorescence and small-angle X-ray scattering, reveal the build-up of two partially folded intermediate conformational states as the pH is decreased (low pH destabilized the protein and accelerated the kinetics of aggregation). A relatively nativelike intermediate, I(N), was observed between pH 4 and 6, with little loss of secondary structure, but with significant tertiary structure changes and enhanced ANS binding, indicating exposed hydrophobic surfaces. At pH below 3, we observed a relatively unfolded, but compact, intermediate, I(U), which was characterized by decreased tertiary and secondary structure. The I(U) intermediate readily forms amyloid fibrils, whereas I(N) preferentially leads to amorphous aggregates. Except at pH 2, where negligible amorphous aggregate is formed, the amorphous aggregates formed significantly more rapidly than the fibrils. This is the first indication that different partially folded intermediates may be responsible for different aggregation pathways (amorphous and fibrillar). The data support the hypothesis that amyloid fibril formation involves the ordered self-assembly of partially folded species that are critical soluble precursors of fibrils.
308 citations
TL;DR: The data suggest that Congo red is sandwiched between two protein molecules causing protein oligomerization, which shows that it must be used with caution as a diagnostic test for the presence of amyloid fibrils in vitro.
307 citations
TL;DR: In photoionization of free, unoriented chiral molecules with circularly polarized radiation, a significant circular dichroism has been observed in the photoelectron angular distribution, which leads to an asymmetry in the momentum transfer to the photoions.
Abstract: In photoionization of free, unoriented chiral molecules with circularly polarized radiation, a significant circular dichroism, ie, an asymmetry in the forward-backward electron emission, has been observed in the photoelectron angular distribution This leads also to an asymmetry in the momentum transfer to the photoions The spectra for the left- and right-handed enantiomers of bromocamphor exhibit asymmetries up to several percent which vary as a function of orbital binding energy This enantioselective effect can similarly occur for biomolecules with handedness, like amino acids, and may thus be a contributing factor related to the origin of the terrestrial biomolecular homochirality
305 citations
TL;DR: Intramolecular folding of a telomeric oligonucleotide into a quadruplex led to fluorescence resonance energy transfer (FRET) between a donor and an acceptor covalently attached to the 5′ and 3′ ends of the DNA, respectively.
Abstract: The secondary structure of guanine-rich oligodeoxynucleotides has been investigated with fluorescent probes. Intramolecular folding of a telomeric oligonucleotide into a quadruplex led to fluorescence resonance energy transfer (FRET) between a donor (fluorescein) and an acceptor (tetramethylrhodamine) covalently attached to the 5' and 3' ends of the DNA, respectively. Depending on oligonucleotide length, quenching efficiency varied between 0.45 and 0.72 at 20 degrees C. The conjugation of the dyes to the oligonucleotide had a limited, but significant, influence on the thermodynamics of G-quartet formation. Intramolecular folding was demonstrated from the concentration independence of fluorescence resonance energy transfer over a wide concentration range. Folding of the oligonucleotide was confirmed by UV absorption, UV melting, and circular dichroism experiments. The folding of the G-quartet could be followed at concentrations as low as 100 pM. Fluorescence resonance energy transfer can thus be used to reveal the formation of multistranded DNA structures.
225 citations
TL;DR: It is observed that a composition of at least 50% alpha-chiral, aromatic residues is necessary for the formation of stable helical structure in hexameric sequences and implications for the design of helical, biomimetic peptoids are discussed.
Abstract: The achiral backbone of oligo-N-substituted glycines or “peptoids” lacks hydrogen-bond donors, effectively preventing formation of the regular, intrachain hydrogen bonds that stabilize peptide α-helical structures. Yet, when peptoids are N-substituted with α-chiral, aromatic side chains, oligomers with as few as five residues form stable, chiral, polyproline-like helices in either organic or aqueous solution. The adoption of chiral secondary structure in peptoid oligomers is primarily driven by the steric influence of these bulky, chiral side chains. Interestingly, peptoid helices of this class exhibit intense circular dichroism (CD) spectra that closely resemble those of peptide α-helices. Here, we have taken advantage of this distinctive spectroscopic signature to investigate sequence-related factors that favor and disfavor stable formation of peptoid helices of this class, through a comparison of more than 30 different heterooligomers with mixed chiral and achiral side chains. For this family of peptoi...
221 citations
TL;DR: The intermolecular aggregation of the oligomers in aqueous solutions revealed a chain length dependent association that required the presence of a stable helical conformation.
Abstract: Circular dichroism spectroscopy has been used to study the self-assembly of two series of m-phenylene ethynylene oligomers in highly polar solvents. The helical conformation of shorter oligomer lengths was found to be stabilized in aqueous acetonitrile solutions, while longer oligomers began to interact intermolecularly. The intermolecular aggregation of the oligomers in aqueous solutions revealed a chain length dependent association that required the presence of a stable helical conformation. Evidence for intermolecular interactions is provided by Sergeants and Soldiers experiments in which the twist sense bias of a chiral oligomer is transferred to an achiral oligomer.
219 citations
TL;DR: A method to stabilize collagen-like peptides by fusing them to the N terminus of the bacteriophage T4 fibritin foldon domain is reported, and it is shown that the foldondomain, which comprises only 27 amino acid residues, forms an obligatory trimer with a high degree of thermal stability.
TL;DR: The structuring preferences of exendin-4 and GLP-1 are determined by NMR in both the solution and dodecylphosphocholine (DPC) micelle-associated states; both peptides display significant helicity from residue 7 to residue 28 with greater fraying at the N-terminus, and secondary structure definition from chemical shift deviations may be the most appropriate treatment for peptides that lack tertiary structure.
Abstract: Exendin-4, a 39 amino acid peptide originally isolated from the oral secretions of the lizard Heloderma suspectum, has been shown to share certain activities with glucagon-like-peptide-1 (GLP-1), a 30 amino acid peptide. We have determined the structuring preferences of exendin-4 and GLP-1 by NMR in both the solution and dodecylphosphocholine (DPC) micelle-associated states. Based on both chemical shift deviations and the pattern of intermediate range NOEs, both peptides display significant helicity from residue 7 to residue 28 with greater fraying at the N-terminus. Thornton and Gorenstein [(1994) Biochemistry 33, 3532-3539] reported that the presence of a flexible, helix-destabilizing, glycine at residue 16 in GLP-1 was an important feature for membrane and receptor binding. Exendin-4 has a helix-favoring glutamate as residue 16. In the micelle-associated state, NMR data indicate that GLP-1 is less helical than exendin-4 due to the presence of Gly16; chemical shift deviations along the peptide sequence suggest that Gly16 serves as an N-cap for a second, more persistent, helix. In 30 vol-% trifluoroethanol (TFE), a single continuous helix is evident in a significant fraction of the GLP-1 conformers present. Exendin-4 has a more regular and less fluxional helix in both media and displays stable tertiary structure in the solution state. In the micelle-bound state of exendin-4, a single helix (residues 11-27) is observed with residues 31-39 completely disordered and undergoing rapid segmental motion. In aqueous fluoroalcohol or aqueous glycol, the Leu21-Pro38 span of exendin-4 forms a compact tertiary fold (the Trp-cage) which shields the side chain of Trp25 from solvent exposure and produces ring current shifts as large as 3 ppm. This tertiary structure is partially populated in water and fully populated in aqueous TFE. The Leu21-Pro38 segment of exendin-4 may be the smallest protein-like folding unit observed to date. When the Trp-cage forms, fraying of the exendin-4 helix occurs exclusively from the N-terminus; backbone NHs for the C-terminal residues of the helix display H/D exchange protection factors as large as 10(5) at 9 degrees C. In contrast, no tertiary structure is evident when exendin-4 binds to DPC micelles. An energetically favorable insertion of the tryptophan ring into the DPC micelle is suggested as the basis for this change. With the exception of exendin-4 in media containing fluoro alcohol cosolvents, NMR structure ensembles generated from the NOE data do not fully reflect the conformational averaging present in these systems. Secondary structure definition from chemical shift deviations may be the most appropriate treatment for peptides that lack tertiary structure.
TL;DR: The results demonstrate that the tetramer to folded monomer transition is insufficient for fibril formation; further tertiary structural changes within the monomer are required.
Abstract: Transthyretin (TTR) is a soluble human plasma protein that can be converted into amyloid by acid-mediated dissociation of the homotetramer into monomers. The pH required for disassembly also results in tertiary structural changes within the monomeric subunits. To understand whether these tertiary structural changes are required for amyloidogenicity, we created the Phe87Met/Leu110Met TTR variant (M-TTR) that is monomeric according to analytical ultracentrifugation and gel filtration analyses and nonamyloidogenic at neutral pH. Results from far- and near-UV circular dichroism spectroscopy, one-dimensional proton NMR spectroscopy, and X-ray crystallography, as well as the ability of M-TTR to form a complex with retinol binding protein, indicate that M-TTR forms a tertiary structure at pH 7 that is very similar if not identical to that found within the tetramer. Reducing the pH results in tertiary structural changes within the M-TTR monomer, rendering it amyloidogenic, demonstrating the requirement for partia...
TL;DR: In this article, the secondary structure of bovine β-casein was re-examined using Fourier transform infrared and circular dichroism spectroscopies to test these predictions.
TL;DR: A cationic peptide with a unique structure and an ability to interact with membranes and to affect intracellular synthesis of proteins, RNA, and DNA is introduced.
TL;DR: Chain length-related factors that stabilize organosoluble peptoid helices of this class are discussed, which are important for the design of helical, biomimetic peptoids sharing this structural motif.
Abstract: Oligomeric N-substituted glycines or “peptoids” with α-chiral, aromatic side chains can adopt stable helices in organic or aqueous solution, despite their lack of backbone chirality and their inability to form intrachain hydrogen bonds. Helical ordering appears to be stabilized by avoidance of steric clash as well as by electrostatic repulsion between backbone carbonyls and π clouds of aromatic rings in the side chains. Interestingly, these peptoid helices exhibit intense circular dichroism (CD) spectra that closely resemble those of peptide α-helices. Here, we have utilized CD to systematically study the effects of oligomer length, concentration, and temperature on the chiral secondary structure of organosoluble peptoid homooligomers ranging from 3 to 20 (R)-N-(1-phenylethyl)glycine (Nrpe) monomers in length. We find that a striking evolution in CD spectral features occurs for Nrpe oligomers between 4 and 12 residues in length, which we attribute to a chain length-dependent population of alternate struct...
TL;DR: The reversibility of the adsorption-desorption cycle was established by comparing the thermostability and secondary structure of BSA before adsorptive, adsorbed on, and exchanged from silica particles with that observed for dissolved BSA that has passed a heating-cooling cycle.
TL;DR: In this article, the effect of binding and conformational changes induced by anionic surfactants such as sodium dodecyl sulfate (SDS) and SOS on bovine serum albumin (BSA) was studied using differential scanning calorimetry (DSC), circular dichroism (CD), fluorescence and UV spectroscopic methods.
Abstract: The effect of binding and conformational changes induced by anionic surfactants sodium dodecyl sulfate (SDS) and sodium octyl sulfate (SOS) on bovine serum albumin (BSA) have been studied using differential scanning calorimetry (DSC), circular dichroism (CD), fluorescence and UV spectroscopic methods. The denaturation temperature, van't Hoff enthalpy and calorimetric enthalpy of BSA in the presence of SDS and SOS and urea at pH 7 have been determined. The results indicate that SDS plays two opposite roles in the folding and stability of BSA. It acts as a structure stabiliser at a low molar concentration ratio of SDS/BSA and as a destabilizer at a higher concentration ratio as a result of binding of SDS to denatured BSA. The Brandts and Lin model has been used to simulate the results.
TL;DR: Hydrophobic modifications at the C-terminus of Shh resulted in only a 2-3-fold increase in activity, and no activation was found with hydrophobic modification at other surface positions, and the degree of activation appears to correlate with the hydrophobicity of the modification rather than any specific chemical feature of the adduct.
Abstract: Post-translational modifications of the developmental signaling protein Sonic hedgehog (Shh) by a long-chain fatty acid at the N-terminus and cholesterol at the C-terminus greatly activate the protein in a cell-based signaling assay. To investigate the structural determinants of this activation phenomenon, hydrophobic and hydrophilic moieties have been introduced by chemical and mutagenic methods to the soluble N-terminal signaling domain of Shh and tested in both in vitro and in vivo assays. A wide variety of hydrophobic modifications increased the potency of Shh when added at the N-terminus of the protein, ranging from long-chain fatty acids to hydrophobic amino acids, with EC50 values from 99 nM for the unmodified protein to 0.6 nM for the myristoylated form. The N-myristoylated Shh was as active as the natural form having both N- and C-terminal modifications. The degree of activation appears to correlate with the hydrophobicity of the modification rather than any specific chemical feature of the adduc...
TL;DR: LD and CD, like other optical techniques, offer possibilities for rapid characterization of nucleic acid complexes using relatively small amounts of sample, but the information that may be gained varies with the level of sophistication at which the measurements are carried out and interpreted.
Abstract: Publisher Summary Polarized light spectroscopy offers possibilities to quickly characterize nucleic acids and their complexes with bound proteins or drug molecules, using a relatively small amount of sample. Linear dichroism (LD) provides structure information in terms of relative orientation between the bound drug molecule and the DNA molecular long axis, and also about the effects of ligand binding on the host. Circular dichroism (CD) may contribute additional structural details about such complexes. LD and CD, like other optical techniques, offer possibilities for rapid characterization of nucleic acid complexes using relatively small amounts of sample. The information that may be gained varies with the level of sophistication at which the measurements are carried out and interpreted. The presence of detectable LD thus, means that the ligand is bound to the oriented DNA, while the sign and amplitude of the LD signal contain information about the nature of the predominant binding mode. Use of complementary techniques, such as CD and fluorescence spectroscopy, may allow further conclusions regarding the structure and dynamics of DNA-ligand complexes. Nonchiral drug molecules (molecules that lack handedness and thereby optical activity) have no CD signal.
TL;DR: The present reconstitution of supramolecular membranes provides the first example of complementary hydrogen-bond-directed formation of soluble, mesoscopic supramolescular assemblies in water.
Abstract: Spontaneous formation of complementary hydrogen-bond pairs and their hierarchical self-assembly (reconstitution) into chiral supramolecular membranes are achieved in water by mixing amphiphilic pairs of glutamate-derived melamine 6 and ammonium-derivatized azobenzene cyanuric acid 4. Electron microscopy is used to observe formation of helical superstructures, which are distinct from the aggregate structures observed for each of the single components in water. In addition, a spectral blue-shift and induced circular dichroism (ICD) with exciton coupling are observed for the π−π* absorption of the azobenzene chromophores. These observations are consistent with the reconstitution of the hydrogen-bond-mediated supramolecular membrane 6−4. Spectral titration experiments indicate the stoichiometric integration of the complementary subunits with an association constant of 1.13 × 105 M-1. This value is considerably larger than those reported for the artificial hydrogen-bonding complexes in aqueous media. The remar...
TL;DR: The schizophyllan complex can be characterized by enhancement of the base stacking, cooperative dissociation, the similar thermodynamic parameters to DNA, and combination of the hydrogen-bonding and hydrophobic interactions to form the higher-order structure.
TL;DR: The results demonstrated that biosensor technology provides a new experimental approach to the study of peptide-membrane interactions through the rapid determination of the binding affinity of bioactive peptides for phospholipids.
TL;DR: The helix-coil transition of a synthetic alpha-helical peptide (the D-Arg peptide), Ac-YGG(KAAAA)(3)-CO-D-Arg-CONH(2), was studied by static far-UV circular dichroism and time-resolved infrared spectroscopy coupled with the laser-induced temperature-jump technique for rapid relaxation initiation.
Abstract: The helix-coil transition of a synthetic alpha-helical peptide (the D-Arg peptide), Ac-YGG(KAAAA)(3)-CO-D-Arg-CONH(2), was studied by static far-UV circular dichroism (CD) and time-resolved infrared spectroscopy coupled with the laser-induced temperature-jump technique for rapid relaxation initiation. Equilibrium thermal unfolding measurements of the D-Arg peptide monitored by CD spectroscopy reveal an apparent two-state helix-coil transition, with a thermal melting temperature around 10 degrees C. Time-resolved infrared (IR) measurements following a laser-induced temperature jump, however, reveal biphasic (or multiphasic) relaxation kinetics. The fast phase rises within the 20 ns response time of the detection system. The slow phase has a decay lifetime of approximately 140 ns at 300 K and exhibits monotonic temperature dependence with an apparent activation energy around 15.5 kcal/mol.
TL;DR: SANS data analysis using the sphere method confirmed the structures imaged by cryo-TEM and provided a detailed structural characterization of amphiphiles that have a model collagen peptide headgroup and showed that the peptide's capacity to organize into a triple helix by intertwining with two neighboring molecules can be affected by increasing the tail length.
Abstract: We have used cryo-transmission electron microscopy (cryo-TEM), small-angle neutron scattering (SANS), differential scanning calorimetry (DSC), and circular dichroism (CD) for microstructural characterization of amphiphiles that have a model collagen peptide headgroup. Single-tail amphiphiles and double-tail amphiphiles with short tails such as C12 and C14 formed spheroidal micelles. Further increase in tail length of the double-tail amphiphiles led to the formation of disklike micelles that aggregated to form a strandlike structure. SANS curves for double-tail amphiphiles were obtained at different contrasts by using different fractions of D2O in the D2O/H2O mixture. SANS data analysis using the sphere method confirmed the structures imaged by cryo-TEM and provided a detailed structural characterization. CD data showed that the peptide's capacity to organize into a triple helix by intertwining with two neighboring molecules can be affected by increasing the tail length. Double-tail amphiphiles with tails ...
TL;DR: The results show that the intrinsically unstructured ER α-N interacts with TBP, and suggest that structural changes are induced in ERα-N upon TBP interaction, which supports models of TBP as a target protein for the N-terminal activation domain of ERα and suggests that target proteins can induce and/or stabilize ordered structure in N- terminal regions of nuclear receptors upon interaction.
TL;DR: These experiments confirm that the thiolate ligand facilitates the O--O bond cleavage by P450 enzymes and also demonstrate that this ligand satisfies important roles in protein folding, substrate binding, and electron transfer.
Abstract: To examine the roles of the proximal thiolate iron ligand, the C357H mutant of P450(cam) (CYP101) was characterized by resonance Raman, UV, circular dichroism, and activity measurements. The C357H mutant must be reconstituted with hemin for activity to be observed. The reconstituted enzyme is a mixture of high and low spin species. Low temperature (10 degrees C), low enzyme concentration (1 microM), high camphor concentration (1 mM), and 5--50 mM buffer concentrations increase the high to low spin ratio, but under no conditions examined was the protein more than 60% high spin. The C357H mutant has a poorer K(m) for camphor (23 vs 2 microM) and a poorer K(d) for putidaredoxin (50 vs 20 microM) than wild-type P450(cam). The mutant also exhibits a greatly decreased camphor oxidation rate, elevated uncoupling rate, and much greater peroxidase activity. Electron transfer from putidaredoxin to the mutant is much slower than to the wild-type even though redox potential measurements show that the electron transfer remains thermodynamically favored. These experiments confirm that the thiolate ligand facilitates the O--O bond cleavage by P450 enzymes and also demonstrate that this ligand satisfies important roles in protein folding, substrate binding, and electron transfer.
TL;DR: In this paper, the effects of temperature and solvents on the helical conformation of poly(propiolic esters) with various chiral side chains, which were prepared with [(nbd)RhCl]2, were investigated using circular dichromism spectroscopy.
Abstract: The effects of temperature and solvents on the helical conformation of stereoregular cis−transoidal poly(propiolic esters) [(CHCCO2R*)n] with various chiral side chains, which were prepared with [(nbd)RhCl]2, were investigated using circular dichromism spectroscopy. CD effects of the polymers with short side chains, where R* = (S)-(CH2)nCH(CH3)C2H5 (n = 1−4), were only slightly amplified with decreasing temperature from +20 to −50 °C, and this behavior was independent of the solvents examined (CHCl3, THF, and n-hexane). In contrast, although the temperature dependence of CD effects for polymers with long alkyl pendants [5 and 6, where R* = (S)-(CH2)5CH(CH3)C2H5 and (S)-(CH2)2CH(CH3)(CH2)3CH(CH3)2, respectively] was similar in n-hexane to those for the other polymers, the Cotton effects of 5 and 6 in CHCl3 decreased in intensity with decreasing temperature and inverted in sign between −30 and −40 °C. This CD inversion process was proved to originate from the thermally induced screw-sense inversion, and com...
TL;DR: The vortex effect can be attributed to enhancement of the chirality fluctuations that originate in the diffusion-limited aggregation to high-molecular-weight homoassociates, and could be general for supramolecular systems that are obtained under kinetic control.
Abstract: Homoassociates of the achiral title porphyrins in acid solutions show spontaneous symmetry breaking, which can be detected by circular dichroism (CD). The CD spectra are due to differential scattering and differential absorption contributions, the relative significance of which is related to the shape and size of the homoassociate. When an earlier model, designed for the association of these diprotonated porphyrins (J aggregates with the geometry of stepped sheets of intramolecular-stabilised zwitterions), was applied to an exciton-coupling point-dipole approximation, the folding of the one-dimensional homoassociates explained the CD signals detected. An effect of the vortex direction, caused by stirring or rotary evaporation, upon the exciton chirality sign was detected. In the case of H2TPPS3, the number of experiments performed gave a statistical significance to this effect. This vortex effect can be attributed to enhancement of the chirality fluctuations that originate in the diffusion-limited aggregation to high-molecular-weight homoassociates. In this sense, the phenomenon could be general for supramolecular systems that are obtained under kinetic control, and its detection would be possible when inherent chiral chromophores were being generated in the association process.