Site frequency spectra from genomic SNP surveys

Genomic survey data now permit an unprecedented level of sensitivity in the detection of departures from canonical evolutionary models, including expansions in population size and selective sweeps. Here, we examine the effects of seemingly subtle differences among sampling distributions on goodness of fit analyses of site frequency spectra constructed from single nucleotide polymorphisms. Conditioning on the observation of exactly two alleles in a random sample results in a site frequency spectrum that is independent of the scaled rate of neutral substitution (θ). Other sampling distributions, including conditioning on a single mutational event in the sample genealogy or randomly selecting a single mutation from a genealogy with multiple mutations, have distinct site frequency spectra that show highly significant departures from the predictions of the biallelic model. Some aspects of data filtering may contribute to significant departures of site frequency spectra from expectation, apart from any violation of the standard neutral model.     

Analysis of multi-component fluorescence emission by phase-sensitive detection using one modulation frequency

We describe the use of phase-sensitive detection of fluorescence to resolve the lifetimes and fractional intensities from multi-component fluorescence samples, using data obtained at a single modulation frequency. Phase-sensitive spectra of the mixture are recorded at arbitrarily chosen detector phase angles. The steady-state spectrum of each component must be known. The phase-sensitive spectra are fitted, using a nonlinear least-squares algorithm, to obtain the lifetimes and fractional intensities of each fluorophore in the mixture. Simulations for two- and three-component mixtures are presented to illustrate how the resolution is affected by spectral overlap and lifetime separation. Experimentally, we resolved two- and three-component mixtures of protein-like fluorophores (N-acetyl-L-tyrosinamide, N-acetyl- L-tryptophanamide, indole and 2,3-dimethylindole) using data collected at 30 MHz. These fluorophores have closely spaced lifetimes of 1.5, 2.9, 4.5 and 4.3 ns, respectively, and display extensive spectral overlap. These results demonstrate that phase-sensitive spectra, recorded at only one modulation frequency with a standard phase fluorometer, can be used to resolve multi-component emissions.     

Spectral fitting for signal assignment and structural analysis of uniformly <Superscript>13</Superscript>C-labeled solid proteins by simulated annealing based on chemical shifts and spin dynamics

We describe an approach for the signal assignment and structural analysis with a suite of two-dimensional (13)C-(13)C magic-angle-spinning solid-state NMR spectra of uniformly (13)C-labeled peptides and proteins. We directly fit the calculated spectra to experimental ones by simulated annealing in restrained molecular dynamics program CNS as a function of atomic coordinates. The spectra are calculated from the conformation dependent chemical shift obtained with SHIFTX and the cross-peak intensities computed for recoupled dipolar interactions. This method was applied to a membrane-bound 14-residue peptide, mastoparan-X. The obtained C', C(alpha) and C(beta) chemical shifts agreed with those reported previously at the precisions of 0.2, 0.7 and 0.4 ppm, respectively. This spectral fitting program also provides backbone dihedral angles with a precision of about 50 degrees from the spectra even with resonance overlaps. The restraints on the angles were improved by applying protein database program TALOS to the obtained chemical shifts. The peptide structure provided by these restraints was consistent with the reported structure at the backbone RMSD of about 1 A.     

Translational and Rotational Diffusion Constants of Tobacco Mosaic Virus from Rayleigh Linewidths

The translational and rotational diffusion constants of tobacco mosaic virus (TMV) have been determined from homodyne and heterodyne measurements of the spectrum of laser light scattered from dilute aqueous solutions of TMV. Our results for the translational and rotational constants respectively, reduced to 20 degrees C, are: D(T) = 0.280 +/- 0.006 x 10(-7) cm(2)/sec, and D(R) = 320 +/- 18 sec(-1). We include a theoretical derivation of the spectrum of light scattered from rod-shaped molecules which reproduces results obtained previously by Pecora, but which is specialized at the outset to the problem of dilute solutions so that simple single-particle correlation functions may be utilized. An analysis of the photocurrent spectrum for both the homodyne and heterodyne detection schemes is given. Various data reduction schemes utilized in the analysis of our spectra are described in some detail, and our results are compared with values of the diffusion constants obtained from other experiments.     

An infrared study of unordered poly-L-proline in CaCl 2 solutions

Infrared spectra have been obtained of poly-L -proline in aqueous CaCl₂ solutions. As the salt concentration is increased, the C?O stretching band develops a component at the frequency found in the solid state while the CH₂ bending band broadens to higher frequency. Since circular dichroism spectra indicate progressive disordering of the chain with increasing salt concentration, we associate the infrared spectral changes with the same phenomenon. Our interpretation of these changes, particularly in the CH₂ bending modes, is that disordering is associated primarily with an increase in the range of accessible C^α–C′ (?O) rotation angles rather than with the random introduction of cis imide bonds in the chain.     

Basal EDRF activity helps to keep the geometrical configuration of arterial bifurcations close to the Murray optimum

We have used X-ray microangiography to investigate the hypothesis that the potent endogenous vasodilator endothelium-derived relaxing factor (EDRF) contributes to the maintenance of "optimality" in vascular branching by modulating the diameters of the parent (D0) and daughter (D1 and D2) arteries at bifurcations. Five anatomically different types of bifurcation were studied in buffer-perfused rabbit ear preparations both under resting conditions and after pharmacological constriction by 5-hydroxytryptamine (5HT). A range of flow rates (1-5 ml min-1) was employed as release of EDRF from endothelial cells is stimulated by shear stress. Experimental data obtained in the presence and absence of EDRF activity were compared with theoretical predictions in three ways. (1) Junction exponents (x) were determined at each bifurcation from the equation Dx1 + Dx2 = Dx0, and their frequency distributions constructed. Murray (1926a, Proc. natn. Acad. Sci., U.S.A. 12, 207-214; 1926b, J. gen. Physiol. 9, 835-841.) proposed that x will be exactly 3 if power losses and intravascular volume are minimized simultaneously. In unconstricted preparations, either in the presence or absence of EDRF activity, and in preparations constricted by 0.1 microM 5HT in the presence of EDRF activity, the modes and medians of the frequency distributions of x were found to be close to 3 at all flow rates. In contrast, in 0.1 microM 5HT-constricted preparations in the absence of EDRF activity, no single mode common to all flow rates was apparent and medians were significantly larger at all flow rates. (2) Theoretically "optimal" branching angles were derived from experimental diameter measurements using four mathematical models which minimize respectively the total surface area, total volume, total drag (shear stress) and total power losses at bifurcations (Murray, 1926b). These calculated branching angles were then compared with actual branching angles. EDRF activity was found to be necessary for accurate prediction of branching angles by the minimum volume and power loss models in 5HT-constricted but not in resting preparations. (3) For each model or "minimization principle", there is an optimal mathematical relationship between the junction exponent, x, and the angle between daughter arteries, psi 12, at a bifurcation (Roy & Woldenberg, 1982, Bull. math. Biol. 44, 349-360.) Experimentally determined values of x and psi 12 agreed closely with those predicted both by the minimum volume and the minimum power loss principles, except again in 5HT-constricted preparations in the absence of EDRF activity.(ABSTRACT TRUNCATED AT 250 WORDS)     

Sound radiation patterns in the frequency domain of cries from a vespertilionid bat

Summary Ultrasonic cries from an immobilized vespertilionid batMyotis daubentoni, were recorded simultaneously in front of the bat (on-axis), and at various off-axis angles. The differences between the on-axis and off-axis spectra were computed and related to the theory of directional emission from a baffled rigid-piston radiator. This theory implies zero radiation at a specific frequency for a given piston diameter and off-axis angle (Fig. 1). The bat's difference spectra showed notches of ca. 25 dB with properties as the zeros of the piston theory (Fig. 7). The structure of off-axis cries was found to be qualitatively predictable from the on-axis cries, using the piston theory (Fig. 4). The difference spectra had a complex, oscillatory fine-structure that could not be accounted for by the basic piston theory.Supported by grants from the Danish Science Research Council for Natural Sciences     

Determination of local conformation of proteins from 1H-NMR spectroscopy data

A method is proposed to determine conformations of amino acid residues of the protein and effective correlation time tau c from cross-peak intensities in two-dimensional nuclear Overhauser enhancement (NOESY) spectra. The method consists in fitting complete relaxation matrix of dipeptide unit protons to experimental cross-peak intensities by varying phi, psi, chi torsional angles and tau c. To verify the method, NOESY spectra of basic pancreatic trypsin inhibitor (BPTI) were theoretically generated at mixing times tau m = 25-300 ms and tau c = 4 ns and used for local structure determination. The method works well with optimum for measurement of NOE intensities tau m 100-200 ms. As a result, the backbone phi, psi torsion angles were unambiguously determined at tau m = 100 ms for all but Gly residues of BPTI, and chi 1 angles were determined for the majority of side chains. The obtained dipeptide unit conformations are very close to the BPTI crystallographic structure: root mean square deviation (RMSD) of interproton distances within dipeptide units, on the average, is 0.08 A (maximal deviation 0.44 A), and RMSD of phi and psi angles are 18 and 9 degrees, respectively (maximal deviations are 44 and 22 degrees).     

Application of constrained regularization analysis to low-angle quasi-elastic light scattering

A constrained regularization procedure has been applied to a low-angle quasi-elastic light scattering system in order to determine particle size distributions. The conditions under which this procedure may be successfully applied to low-angle photon correlation spectroscopy have been characterized. Acquisition of photon count data over a short time period, relative to the long exponential decay constants of correlation functions obtained at low forward angles, resulted in particle size distributions which were stable with regard to peak width and weighted mean particle radius. Irrespective of the number of photon counts obtained, peak resolution and position on the particle size scale were not optimized unless anomalies in the correlation function due to transient increases in the mean photon counting rate were removed from the photon count data prior to autocorrelation. When such measures were taken, reasonable size distributions were obtained for well characterized protein standards and for liposomal suspensions.     

Optical spectroscopic elucidation of beta-turns in disulfide bridged cyclic tetrapeptides

Vibrational circular dichroism (VCD) spectroscopic features of type II beta-turns were characterized previously, but, criteria for differentiation between beta-turn types had not been established yet. Model tetrapeptides, cyclized through a disulfide bridge, were designed on the basis of previous experimental results and the observed incidence of amino acid residues in the i + 1 and i + 2 positions in beta-turns, to determine the features of VCD spectra of type I and II beta-turns. The results were correlated with electronic circular dichroism (ECD) spectra and VCD spectra calculated from conformational data obtained by molecular dynamics (MD) simulations. All cyclic tetrapeptides yielded VCD signals with a higher frequency negative and a lower frequency positive couplet with negative lobes overlapping. MD simulations confirmed the conformational homogeneity of these peptides in solution. Comparison with ECD spectroscopy, MD, and quantum chemical calculation results suggested that the low frequency component of VCD spectra originating from the tertiary amide vibrations could be used to distinguish between types of beta-turn structures. On the basis of this observation, VCD spectroscopic features of type II and VIII beta-turns and ECD spectroscopic properties of a type VIII beta-turn were suggested. The need for independent experimental as well as theoretical investigations to obtain decisive conformational information was recognized.     

Resonance Raman studies of Escherichia coli sulfite reductase hemoprotein. 2. Fe4S4 cluster vibrational modes

Resonance Raman (RR) spectra from the hemoprotein subunit of Escherichia coli sulfite reductase (SiR-HP) are examined in the low-frequency (200-500 cm-1) region where Fe-S stretching modes are expected. In spectra obtained with excitation in the siroheme Soret or Q bands, this region is dominated by siroheme modes. Modes assignable to the Fe4S4 cluster are selectively enhanced, however, with excitation at 488.0 or 457.9 nm. The assignments are confirmed by observation of the expected frequency shifts in SiR-HP extracted from E. coli grown on 34S-labeled sulfate. The mode frequencies and isotopic shifts resemble those seen in RR spectra of other Fe4S4 proteins and analogues, but the breathing mode of the cluster at 342 cm-1 is higher than that observed in the other species. Spectra of various ligand complexes of SiR-HP reveal only slight sensitivity of the cluster terminal ligand modes to the presence of exogenous heme ligands, at variance with a model of ligand binding in a bridged mode between heme and cluster. Close examination of RR spectra obtained with siroheme Soret-band excitation reveals additional 34S-sensitive features at 352 and 393 cm-1. These may be attributed to a bridging thiolate ligand.     

A new method for resolution of two- and three-component mixtures of fluorophores by phase-sensitive detection of fluorescence

We describe a new method for the analysis of phase-sensitive fluorescence emission spectra. This method permits the resolution of three-component mixtures using spectra measured at a single modulation frequency. Phase-sensitive spectra are recorded using one modulation frequency, at a number of arbitrary detector phase angles. It is not necessary to suppress any one component. The spectra are then used to estimate the component lifetimes and steady-state fractional intensities using a nonlinear least-squares analysis procedure. The only requirement for the analysis is the knowledge of the steady-state spectra of the individual components. This procedure allowed the resolution of a two-component mixture of 9-methylanthracene (4.5 ns) and 9,10-diphenylanthracene (5.9 ns). It should be noted that resolution of two lifetimes which differ by only 30% is a difficult task. Additionally, we resolved a three-component mixture with lifetimes that differed fourfold: p-bis[2-(5-phenyloxazolyl)]benzene (1.3 ns), 9-methylanthracene (4.5 ns), and 9,10-diphenylanthracene (5.9 ns). Conveniently, the technique utilizes a commercially available fixed-frequency phase fluorometer.     

Characterization by 1H NMR of glycosidic conformations in the tetramolecular complex formed by d(GGTTTTTGG).

We have conducted two dimensional NOESY studies on the molecule d(G2T5G2) to characterize the structure of the tetramolecular complex previously identified by calorimetric and spectroscopic studies (1). Analysis of the NOE and exchange cross peaks observed in the NOESY spectra establishes the formation of structured conformations at low temperature (5 degrees C). Significantly, within each strand of these structured conformations, the G1 and G8 residues adopt syn glycosidic torsion angles, while the G2 and G9 residues adopt anti glycosidic torsion angles. Consequently, any structure proposed for the tetramolecular complex of d(G2T5G2) must have alternating G(syn) and G(anti) glycosidic torsion angles within each strand. The implications of this observation for potential structures of the tetramolecular complex of d(G2T5G2) are discussed.     

Electromyographic responses to prescribed mastication.

The aim was to understand between-volunteer differences in Electromyography (EMG) behaviour during chewing. EMG was used to record the electrical activity of the temporal and masseter muscles of volunteers, who carried out mastication movements by operating calibrated springs held between their incisors. The volunteers coordinated their jaw movements with the signal produced by a metronome, at four rates: 30, 60, 90 and 120 beats per minute (bpm). Raw data were analyzed to examine the distributions of the intervals between chews. For the highest prescribed chew rates, the volunteers' distributions were very similar. The distributions varied most for the 30 bpm data, suggesting that volunteers differed in their ability to carry out and maintain this prescribed chewing pattern. The data were Fourier transformed to give power spectra in the frequency domain. The low frequency (<10 Hz) region contained spectral features related to the prescribed chew rate. Principal component analysis of the power spectra revealed that readings from each volunteer clustered together, and the clusters could be largely separated. Such grouping was found irrespective of whether data from each chew rate were analyzed separately or simultaneously. This indicated that within-volunteer variance, arising from the different chew rates as well as between-session variance, is lower than between-volunteer variance; even when individuals are asked to make jaw movements in the same prescribed manner, they can nevertheless be uniquely distinguished by their muscle activity as recorded by EMG.     

Simulations of oligopeptide vibrational CD: effects of isotopic labeling

Simulated ir absorption and vibrational CD (VCD) spectra of four alanine-based octapeptides, each having its main chain constrained to a different secondary structure conformation, were analyzed and compared with experimental results for several different peptides. The octapeptide simulations were based on transfer of property tensors from a series of ab initio calculations for a short L-alanine based segment containing 3 peptide bonds with relative straight phi, psi angles fixed to those appropriate for alpha-helix, 3(10)-helix, ProII-like helix, and beta-sheet-like strand. The tripeptide force field (FF) and atomic polar tensors were obtained with density functional theory techniques at the BPW91/6-31G** level and the atomic axial tensor at the mixed BPW91/6-31G**/HF/6-31G level. Allowing for frequency correction due to the FF limitations, the octapeptide results obtained are qualitatively consistent with experimental observations for ir and VCD spectra of polypeptides and oligopeptides in established conformations. In all cases, the correct VCD sign patterns for the amide I and II bands were predicted, but the intensities did have some variation from the experimental patterns. Predicted VCD changes upon deuteration of either the peptide or side-chains as well as for (13)C isotopic labeling of the amide C=O at specific sites in the peptide chain were computed for analysis of experimental observations. A combination of theoretical modeling with experimental data for labeled compounds leads both to enhanced resolution of component transitions and added conformational applicability of the VCD spectra.     

Orientational preferences shown by microspikes of growing nerve cells in vitro

The shapes of microspikes on single neurones cultured in vitro have been analysed with respect to angles of bending and branching. Characteristic frequency distributions were found in all of the four categories of angles. Certain `preferred' orientations of bending and branching were seen to center about 60°, 90°, and 120°. The possible cellular basis for such behavior is discussed, as well as the similarity of bending and branching in the axonal-microspike system of the single nerve cell to the analogous branching in the vertebrate blood vascular system.     

An analysis of the amide I mode in the vibrational spectra of unordered polypeptides and proteins

The difference in the observed frequency of the amide I mode between the Raman and ir spectra of many unordered polypeptides is interpreted in terms of a model in which sequences of amide groups have similar ?, ψ angles. The splittings due to intramolecular interactions for the infinite helices generated by all ?, ψ angles are calculated by assuming transition dipole coupling. The effect of finite sequence length and distortions are then considered. This hypothesis accounts for the general features of the amide I mode of unordered polypeptides.     

Circadian and Ultradian Rhythms in Heart Rate and Motor Activity of Smokers,Abstinent Smokers,and Nonsmokers

In a field study, heart rate and motor activity were assessed continuously in 12 male smokers during 2 smoking and 2 abstinence days and in 12 male nonsmokers during 4 days. A circadian analysis revealed earlier activity acrophases in smokers than nonsmokers and earlier heart rate acrophases in abstinent than smoking smokers. Furthermore, heart rate acrophases of smoking smokers significantly anticipated activity acrophases;, whereas in abstinent smokers and nonsmokers the two parameters oscillated in phase. With the use, in smoking smokers, of the individual average smoking interval as a hypothetical ultradian period length, significant periodicities were found for heart rates in 16 and for activity in 15 of 24 observation days. These rhythms were nicotine independent and based on heart rate and activity increases prior to lighting up the cigarettes. Individual frequency spectra for the 16 h after getting up and the 7 h after going to bed did not reveal single dominant frequencies but rather complex frequency distributions. Power spectra of the daytime data revealed no group differences for activity and no heart rate differences between smoking smokers and nonsmokers. In abstinent smokers, however, a significant reduction of heart rate frequencies slower than 1 cycle/135 min and a significant increase of heart rate frequencies faster than 1 cycle/20 min were observed as compared with all other groups. This effect persisted over the 2 abstinence days, suggesting an activity-independent change in the frequency distribution of heart rates after quitting smoking.     

Circadian and Ultradian Rhythms in Heart Rate and Motor Activity of Smokers, Abstinent Smokers, and Nonsmokers

In a field study, heart rate and motor activity were assessed continuously in 12 male smokers during 2 smoking and 2 abstinence days and in 12 male nonsmokers during 4 days. A circadian analysis revealed earlier activity acrophases in smokers than nonsmokers and earlier heart rate acrophases in abstinent than smoking smokers. Furthermore, heart rate acrophases of smoking smokers significantly anticipated activity acrophases;, whereas in abstinent smokers and nonsmokers the two parameters oscillated in phase. With the use, in smoking smokers, of the individual average smoking interval as a hypothetical ultradian period length, significant periodicities were found for heart rates in 16 and for activity in 15 of 24 observation days. These rhythms were nicotine independent and based on heart rate and activity increases prior to lighting up the cigarettes. Individual frequency spectra for the 16 h after getting up and the 7 h after going to bed did not reveal single dominant frequencies but rather complex frequency distributions. Power spectra of the daytime data revealed no group differences for activity and no heart rate differences between smoking smokers and nonsmokers. In abstinent smokers, however, a significant reduction of heart rate frequencies slower than 1 cycle/135 min and a significant increase of heart rate frequencies faster than 1 cycle/20 min were observed as compared with all other groups. This effect persisted over the 2 abstinence days, suggesting an activity-independent change in the frequency distribution of heart rates after quitting smoking.