Theoretical π-π* absorption and circular dichroic spectra of polypeptide β-structures

Absorption and circular dichroic (CD) spectra of the π-π* transition near 200 nm are calculated for poly(Gly), poly(Ala), poly(Abu), and poly(Val) in the β_P (paralle) and β_A (antiparallel) pleated-sheet structures using the dipole interaction model and including interactions among all atoms; optical parameters were obtained from previous studies of related molecules. Most of the calculations are for structures with one or three chains of six residues each. The oscillator strength and splitting of the π-π* modes are found to be affected only to a small extent by variations in side-chain structure and conformation, whereas the CD spectrum is very sensitive to these variations. Poly(Gly) and poly(Ala) β-structures in uniform, planar lattices do not show sufficient rotational strength to account for observed CD spectra. Poly(Abu) and poly(Val) β-structures in uniform, planar lattices show rotational strengths comparable to experiment when χ¹ is near ?60° for β_A-structures or in a broad range near 140° for β_P-structures. Poly(Ala) in uniformly twisted β_A- and β_P-structures or in irregular β_A-structures corresponding to regions of the crystal structure of concanavalin A also show enhanced rotational strengths in the principal π-π* CD band. Absorption and CD spectra calculated for poly(Abu) in uniform β_A- and β_P-structures are compared with experimental data on poly(Lys) in the β-form, assuming side-chain conformations in Abu that maximize the intensity of the principal CD band. The calculations for the β_A-form show the better agreement with experiment for both types of spectra.     

Effects of proline ring conformation on theoretical pi-pi* absorption and CD spectra of helical poly(L-proline) forms I and II

Absorption and CD spectra of the pi-pi* transition near 200 nm are calculated for helical (Pro)10 forms I and II with a variable proline ring conformation characterized by torsion angle chi 2 in the range -60 degrees to 60 degrees. The spectra for poly(Pro) I are not sufficiently sensitive to chi 2 to suggest a preferred ring conformation. The spectra for poly(Pro) II are more sensitive to chi 2, and suggest preferred ring conformations near either or both of the chi 2 regions -50 +/- 10 degrees and 50 +/- 10 degrees.     

Reassessment of the random coil conformation: vibrational CD study of proline oligopeptides and related polypeptides.

The "random coil" conformational problem is examined by comparison of vibrational CD (VCD) spectra of various polypeptide model systems with that of proline oligomers [(Pro)n] and poly(L-proline). VCD, ir and uv CD spectra of blocked L-proline oligopeptides [(Pro)n, n = 2-12] in different solvents are reported and compared to the spectra of poly(L-proline) II, poly(L-glutamic acid), and unblocked proline oligomers. Based on the chain-length dependence of the VCD and electronic CD (ECD) spectra of proline oligomers, it is established that VCD spectra are dominated by short-range interactions. The VCD of random coil model polypeptides is shown to be identical in shape but smaller in magnitude than poly(L-proline) II and of similar magnitude to that of (Pro)n (n = 3, 4). Based on the spectral evidence, it is concluded that the "random coil" conformation has a large fraction of helical regions, conformationally similar to the left-handed, 3(1) polyproline II helix, as was previously suggested by Krimm and co-workers. This conclusion is further supported by studies of effects of salt (CaCl2, LiBr, LiClO4), temperature (5-75 degrees C), and pH on the VCD spectra of L-proline oligomers, poly(L-proline) II, and poly(L-glutamic acid). These show that, after each of these perturbations, a significant local ordering remains in the oligomers and polymers studied, and that charged polypeptides such as poly(L-glutamic acid) are more flexible than are polyproline or even L-proline oligomers.     

Induced CD of DNA intercalators: electric dipole allowed transitions

The induced CD of an electric dipole allowed transition of a DNA intercalator has been calculated using the “matrix method” and a set of DNA ππ* transitions recently adopted for calculating the CD of DNA by Rizzo and Schellman [(1984) Biopolymers 23 , 435–470]. The induced CD is strongly dependent on the angular orientation of the intercalator and only moderately dependent on its location within the intercalation pocket. The dependence of the CD on the orientation and location of the intercalator was studied for some representative conformations of di- and tetranucleotide duplexes of (dGdC) and (dAdT). The effect of alternative DNA transition moment directions was also tested. The orientation dependence compares well with the previously predicted 1-2 cos² γ dependence [B. Nordén and F. Tjerneld (1982) Biopolymers 21 , 1713–1734]. Measured induced CD spectra of methylene blue (MB) intercalated in double-stranded poly(dAdT), poly(dGdC), and calf-thymus DNA are discussed on the basis of the results of the calculations. Major differences between the induced CD spectra are likely to reflect different modes of intercalation for the different nucleotide sequences. In particular, the concluded geometry in solution for MB intercalated in poly(dAdT) differs significantly from the corresponding geometry found in dinucleotide–intercalator crystals.     

Circular dichroism of nucleic acid monomers. II. Derivation and application to polymers of a consistent set of guanine and cytosine transition parameters

An approximate semiempirical procedure has been developed in order to derive nucleic acid monomer π → π* electronic transition moment parameters. Using the approximate procedure, guanine (G) and cytosine (C) transition moment parameters have been derived from agreement found between calculated weight-averaged and measured CD spectra of cyclic-GMP and cyclic-CMP. The derived base transition moment parameters have been assessed in CD spectral calculations on some G- and C-containing nucleic acids for which reasonably good structural information exists. An attempt was also made at evaluating the likely CD spectral contributions of G and C electric n → π* transition moments whose magnitudes were taken to be the maximum expected. Overall, the results indicate that the derived G and C π → π* transition moment parameters are more successful in nucleic acid CD spectral calculations than those used in previous DeVoe theory CD calculations. In addition, the results indicate that electric n → π* transitions may be of importance in understanding nucleic acid monomer CD spectra but appear to be relatively unimportant in understanding nucleic acid polymer CD spectra. It is concluded that the derived G and C π → π* parameters are more useful in DeVoe theory CD calculations than parameters used previously.     

Unusual CD couplet pattern observed for the pi*<--n transition of enantiopure (Z)-8-methoxy-4-cyclooctenone: an experimental and theoretical study by electronic and vibrational circular dichroism spectroscopy and density functional theory calculation

Ultraviolet absorption (UV) and electronic circular dichroism (ECD) spectra of enantiopure (Z)-8-methoxy-4-cyclooctenone (MCO) were measured in hexane to give a normal single UV absorption band at 298 nm, which is assigned to the carbonyl's pi*<--n transition. Unexpectedly, the ECD spectrum exhibited an apparent couplet pattern with vibrational fine structures. Obviously, the conventional CD exciton coupling mechanism cannot be applied to this bisignate CD signal observed for single-chromophoric MCO. Variable temperature-ECD and vibrational circular dichroism (VCD) spectral measurements, simultaneous UV and ECD spectral band resolution, and density functional theory (DFT) calculations of energy and structure revealed that this apparent CD couplet originates from a rather complicated spectral overlap of more than three conformers of MCO, two of which exhibit mirror-imaged ECD spectra at appreciably deviated wavelengths. In the simultaneous band-resolution analysis, the observed UV and ECD spectra were best fitted to four overlapping bands. Two major conformers were identified by comparing the experimental IR and VCD spectra with the simulated ones, and the other two by comparing the observed UV and ECD spectra with the theoretical ones obtained by time-dependent DFT calculations. It was shown that the combined use of experimental ECD and VCD spectra and theoretical DFT calculations can give a reasonable interpretation for the Cotton effects of the conformationally flexible molecule MCO.     

Circular dichroism calculations for double-stranded polynucleotides of repeating sequence

Optical property calculations are presented for poly(A·U), poly[(A-U)·(A-U)], poly(G·C), and poly[(G-C)·(G-C)] in RNA, B-DNA, and C-DNA conformations. An all-order classical coupled oscillator polarizability theory was used, and an effective dielectric constant of 2 was assumed. The calculated CD spectra were found to be sensitive to both geometry and sequence. Agreement with the measured CD spectra of poly(A·U), poly(G·C), and poly(dG·dC) is very good. Calculations for other sequences and geometries are less satisfactory and are particularly poor for poly[(G-C)·(G-C)] in RNA geometry and poly(A·T) in B-DNA geometry. Attempts to improve agreement with measured spectra by varying monomer properties have been only partially successful for these calculations, but they illustrate the types of changes that may prove to be necessary. Calculations using other published X-ray coordinates for certain deoxypolynucleotides of simple sequence, some of which are quite different from B-DNA coordinates, did not result in better agreement with measured spectra. Finally, the dependence of the calculated CD on chain length is examined. Results show that non-nearest neighbor interactions can be important when runs of 3 or more identical base pairs appear in a given sequence.     

Unusual effect of salts on the homodimeric structure of NADH oxidase from Thermus thermophilus in acidic pH

The unusual salt-dependent behavior of the homodimeric flavoenzyme NADH oxidase from Thermus thermophilus in acidic pH has been studied using circular dichroism (CD) and sedimentation velocity. The native-like secondary and quaternary structures in acidic low ionic strength conditions were significantly perturbed by the addition of salts. The peptide region of the CD spectra showed a major salt-induced conformational change in the protein secondary structure. Sedimentation velocity experiments showed dissociation of the homodimeric structure of NADH oxidase in the presence of salt (>1 M). The new acidic conformation of the protein was stabilized by high ionic strength as indicated by a salt-induced increase in the melting temperature of the protein, and by a shift in the apparent pK(a) values of the conformational transition to a less acidic pH. Distortion of the dominant alpha-helical signal was expressed as the disappearance of the parallel polarized Moffitt exciton band at 208 nm without an accompanying loss of amplitude of n-->pi* electronic transitions at 222 nm. The unusual CD spectra correlated qualitatively with the theoretically calculated CD spectra of short alpha-helical structures and/or twisted beta-sheets. Differences between the experimentally obtained CD spectra and theoretical calculations (AGADIR) of the alpha-helical content of NADH oxidase indicate a role for non-local interactions in the protein conformation at high ionic strength and low pH. These findings indicate the importance of the homodimeric interface and electrostatic interactions for maintaining the structural integrity of this thermophilic protein.     

Circular dichroism of collagen, gelatin, and poly(proline) II in the vacuum ultraviolet.

Circular dichroism spectra for acid-soluble calfskin collagen, gelatin, and poly(proline) II in solution have been extended into the vacuum ultraviolet region. The extended spectrum of gelatin reveals that the circular dichroism of this unordered polymer is more closely related to the spectrum of charged polypeptides than might be evident from near ultraviolet work. A short-wavelength band is found at about 172 nm, which corresponds in position, magnitude, and sign to a band recorded earlier for poly(L -glutamic acid) at pH 8.0. This band is observed in a helical structure for the first time in the vacuum ultraviolet circular dichroism and absorption spectra of poly(proline) II. Both circular dichroism and absorption spectra point to the assignement of this band as the nσ*. Neither the nσ* nor the expected positive lobe of the ππ* helix band is observed in the extended circular dichroism spectrum of collagen. We postulate that these two bands cancel here in analogy to the case of α-helical poly(L -glutamic acid).     

Osmotic flow equations for leaky porous membranes

A basic set of equations describing the flows of volume (Jv) and solute (Js) across a leaky porous membrane, coupled to the differences of osmotic and hydrostatic pressures d pi and dP has been derived by using general frictional theory. Denoting the mean pore concentration of solute by c*s and the hydraulic and diffusive conductances by Lp and Ps/RT the equations take the form Jv = LpdP + sigma sLp d pi Js = c*s(1 - sigma f)Jv + Ps d pi/RT sigma s = theta (1 - DsVs/DwVw - Ds/Dos) sigma f = 1 - theta DsVs/DwVw - Ds/Dos in which Dw and Ds are the diffusion coefficients for water and solute in the pore and Dos that for free solution. The relation between the reflection coefficients sigma s and sigma f for osmosis and ultrafiltration is then given by sigma s = sigma f - (1- theta)(1 - Ds/Dos), where theta is the diffusive-driven:pressure-driven flow ratio. These equations follow from the fact that in leaky pores osmosis occurs by diffusion alone and that there cannot be any Onsager symmetry leading to sigma s = sigma f. Symmetry holds in the limits where either the pore is small, when sigma s = sigma f = 1, or where the pore is large when sigma s = sigma f = 0.     

CD of nucleic acids: III. Calculated CD of RNAs from new A, U, G, and C transition-moment parameters

New adenine (A) and uracil (U) π → π* transition-moment parameters have been derived from a recently developed semiempirical procedure. Using conformational energy probabilities based on the Boltzmann equation, the new parameters were assigned by optimizing the calculated CD of cyclic nucleotides against measured CD. The derived A-and U-parameters (along with guanine and cytosine parameters derived previously by the same procedure) have been assessed in CD spectral calculations of some polyribonucleic acid sequences, in assumed A-class geometries. Comparisons have been made between CD spectra calculated from the newly derived parameters and those calculated from parameters obtained from a combination of crystal optical measurements and quantum-mechanical calculations. Although some spectral differences do occur, for the RNA sequences considered, no major disagreements were found in CD spectral signs and shapes, between measurements and calculations. Overall, the results indicate that the newly derived A-, U-, G-, and C-parameters show better agreement between theory and experiment than those used in previous nucleic acid CD calculations.     

Frontier molecular orbital analysis of Cu(n)-O(2) reactivity

Frontier molecular orbital (FMO) theory coupled with density functional calculations has been applied to investigate the chemical reactivity of three key bioinorganic Cu(n)-O(2) complexes, the mononuclear end-on hydroperoxo-Cu(II), the side-on bridged mu-eta(2):eta(2)-O(2)(2-) Cu(II)(2) dimer and the bis-mu-oxo Cu(III)(2) dimer. Two acceptor orbitals (sigma* and pi*) of each complex and two types of donating substrates (sigma-substrate, phosphine; pi-substrate, alkylbenzene) are considered in the electrophilic attack mechanism. The angular dependences of different reaction pathways are determined using FMO theory and the angular overlap model. Including steric effects, the sigma*/sigma and pi*/pi pathways are found more reactive than the corresponding cross sigma*/pi and pi*/sigma pathways which have poor donor-acceptor orbital overlaps in the sterically constrained substrate access region.     

Persistent survival of prevalent clonotypes within an immunodominant HIV gag-specific CD8+ T cell response

CD8(+) T cells play a significant role in the control of HIV replication, yet the associated qualitative and quantitative factors that determine the outcome of infection remain obscure. In this study, we examined Ag-specific CD8(+) TCR repertoires longitudinally in a cohort of HLA-B*2705(+) long-term nonprogressors with chronic HIV-1 infection using a combination of molecular clonotype analysis and polychromatic flow cytometry. In each case, CD8(+) T cell populations specific for the immunodominant p24 Gag epitope KRWIILGLNK (KK10; residues 263-272) and naturally occurring variants thereof, restricted by HLA-B*2705, were studied at multiple time points; in addition, comparative data were collected for CD8(+) T cell populations specific for the CMV pp65 epitope NLVPMVATV (NV9; residues 495-503), restricted by HLA-A*0201. Dominant KK10-specific clonotypes persisted for several years and exhibited greater stability than their contemporaneous NV9-specific counterparts. Furthermore, these dominant KK10-specific clonotypes exhibited cross-reactivity with antigenic variants and expressed significantly higher levels of CD127 (IL-7Rα) and Bcl-2. Of note, we also found evidence that promiscuous TCR α-chain pairing associated with alterations in fine specificity for KK10 variants could contribute to TCR β-chain prevalence. Taken together, these data suggest that an antiapoptotic phenotype and the ability to cross-recognize variant epitopes contribute to clonotype longevity and selection within the peripheral memory T cell pool in the presence of persistent infection with a genetically unstable virus.     

Interaction of amphipathic polypeptides with phospholipids: characterization of conformations and the CD of oriented beta-sheets

The effects of interaction with phospholipids on the conformation of the alternating copolymer, poly(Leu-Lys), and the random copolymer poly(Leu⁵⁰, Lys⁵⁰) have been investigated by CD and ir spectroscopy. Poly(Leu-Lys) undergoes a partial unordered → β-sheet transition in solution in the presence of lysolecithin. On addition of lysolecithin plus cholate, an unordered → α -helix transition is observed. In films deposited from these solutions, poly(Leu-Lys) adopts the anti-parallel β-sheet conformation, as in aqueous solutions at moderate ionic strength. Polarized ir spectra showed that the plane of the β-sheet in such films deviates from the plane of the film by no more than 14°. The random copolymer, poly(Leu⁵⁰, Lys⁵⁰), is α-helical in the presence of lysolecithin and lysolecithin plus cholate, regardless of whether the sample is a solution or a film. CD measurements on the poly(Leu-Lys) films provide information about the component of the CD tensor for light propagating normal to the plane of the β-sheet. These measurements show (1) a negative n → π* CD band (214 nm maximum) with higher intensity than the average CD for isotropic solution; and (2) a positive band in the π → π* region (195 nm maximum), which is weaker than that in the isotropic spectrum.     

Peptide-sandwiched protoporphyrin compounds mimicking hemoprotein structures in solution

A series of covalently bound peptide-protoporphyrin-peptide compounds, also carrying naphthalene (N) to allow a photophysical investigation, were synthesized. Their general formula is P(nN)(2), where P refers to protoporphyrin IX, and n to the number of amino acids in the sequence Boc-Leu-Leu-Lys-(Ala)(x) -Leu-Leu-Lys-OtBu of each backbone chain (x = 0-3; n = x + 6). Their structural features in methanol solution were investigated by ir and CD spectra, and by steady-state and time resolved fluorescence experiments as well. The ir spectra indicate that intramolecularly H-bonded conformations form, and CD data in both methanol and water-methanol mixture suggest the presence of alpha-helix structure. Quenching of excited naphthalene takes place by electronic energy transfer from singlet N* to P ground state. Fluorescence decays coupled with molecular mechanics calculations indicate that two conformers for each dimeric peptide are the major contributors to the observed phenomena. These conformers are characterized by a globular, protein-like structure, where the protoporphyrin resides in a central pocket, while the two N groups are externally situated. Of the four N linkages in the two conformers, three of them attain a very similar steric arrangement around the central P molecule, in terms of both center-to-center distance and mutual orientation, while the fourth experiences a different steric disposition as compared to the others. Experimental photophysical parameters satisfactorily compare with those obtained by theoretical calculations, within the F?rster mechanism for long-range energy transfer, only when the mutual orientation of the chromophores was also taken into account. This implies that interconversion among conformational substates of probes linkages is slow on the time scale of the energy transfer process.     

Matrix-method calculation of linear and circular dichroism spectra of nucleic acids and polynucleotides

Calculations of the optical properties (absorption, linear dichroism, circular dichroism, and anisotropic components of the CD) are presented for polynucleotides of random or regular sequence within the formalism of the matrix method using a set of parameters that includes only the ππ* transitions of the aromatic bases. Experimental solution spectra agree favorably with calculated CD spectra for A-RNA, A-DNA, and B-DNA, when coordinates derived from x-ray studies on fibers are used. Excessive hypochromicity is predicted when parameters intended to reproduce the vacuum-uv absorption of the chromophores are included in the calculations, but total elimination of these parameters leads to an insufficient hypochromicity for the long-wavelength absorption band. Using alternative conformations for DNA in low-salt aqueous solution did not improve the agreement between experimental and calculated spectra, but some features of the optical properties predicted for these variant structures suggest that the tilt of the bases with respect to the helical axis may be larger than that of the fiber B-form. In the case of polynucleotides with regular structure, which have been traditionally less easy to understand in terms of the standard nucleic acid conformations, a series of alternative structures has been examined. Unexpectedly, the calculated spectrum for the Z-DNA structure compares almost quantitatively with the experimental spectrum of poly(dGC·dGC) in low salt. This result, which confirms a recent report [Vasmel, H. & Greve, J. (1981) Biopolymers 20 , 1329–1332], is in contrast with the current identification of Z-DNA with the high-salt form of poly(dGC·dGC). Finally, the optical properties of single-stranded polyribonucleotides appear to be better explained when alternative structures [9₁-helix for poly(rA) and 6₁-helix for poly(rC)] are introduced instead of the A-RNA form.     

Circular dichroism of adenine and thymine containing synthetic polynucleotides

Circular dichroism (CD) spectra of poly(dA), poly(dT), poly(dA)·poly(dT), and poly[d(A-T)]·poly[d(T-A)] have been measured as a function of temperature. From these data difference spectra have been calculated by subtracting the spectrum measured at low temperature from the spectra measured at higher temperatures. The CD difference spectra obtained upon melting of the two double-stranded polymers are very similar. From a comparison of these difference spectra with calculated ones it is shown that optical transitions near 272 nm (on A) and 288 nm (most probably on T) are present. The premelting changes of the CD spectrum of poly[d(A-t)]·poly[d(T-A)] are due to a change in conformation in which the secondary structure goes from a C- to B-type spectrum by increasing the A-type nature of the polymer. Such a change is not observed for poly(dA)·poly(dT). Instead, a transition between two different B-type geometries occurs.     

Theoretical pi-pi absorption, circular dichroic, and linear dichroic spectra of collagen triple helices

Absorption, CD, and LD spectra of the π-π* transition near 200 nm are calculated for poly(Gly-X-Y) (X,Y = Gly, Ala, Pro) in four conformations proposed for collagen like triple helices in the recent literature. A dipole interaction model is used with the same optical paramenters as in previous studies of polypeptide spectra. The CD spectra are sensitive to backbone structure and amino acid composition, although the experimentally observed negative peak near 200 nm is a general feature of most the calculated spectra. Interchain interactions significantly affect the CD spectra in most cases. Calculations for (Gly-Pro-Ala)₃ and (Gly-Ala-Pro)₃ in the triple helical structure of Fraser, MacRae, and Suzuki [(1979) J. Mol. Biol. 129 , 463–481] show absorption, CD, and LD spectra in fairly good agreement with experiment. The characteristics of the π-π* normal modes responsible for the calculated spectra are compared with those of the component bands resolved from the experimental spectra of collagen by Mandel and Holzwarth [(1973) Biopolymers 12 , 655–674].