Loop length, intramolecular diffusion and protein folding

Intramolecular diffusion plays a role in protein folding as shown by kinetic experiments on two alpha-spectrin SH3 domain circular permutants (S19-P20s and N47-D48s), with different poly-glycine loop lengths. Insertion of up to 10 Gly residues does not alter the structure of the folded state nor the overall characteristics of the denatured ensemble. The apparent level of the energy barrier between the denatured and folded species increases linearly with the number of inserted glycines. This suggests that the transition state itself and/or possibly previous transient unstable intermediates are accessed with more difficulty when loop length is increased. The fact that the induced impediment is directly proportional to the number of Gly residues and not to the free energy difference in the folded state indicates that diffusion of different parts of the molecule relative to each other is taking place on going from the denatured ensemble to the transition state. Our results also suggest that transition state ensembles could be more homogenous than recently postulated.     

Simulation of the elementary damage accumulation kinetics in loaded materials

The evolution of an ensemble of elementary-damage clusters is considered using a probabilistic cellular automaton, and the characteristics of the time series of the number of elementary damages and the number of elementary-damage clusters are studied. The transition of the autocorrelation function of the time series of the number of elementary damages to a negative-correlation region is shown to correspond to the transition of the evolution of an ensemble of clusters to the stage of coalescence. The autocorrelation function of the time series of the number of elementary damages agrees qualitatively with the experimental behavior of the autocorrelation functions of the number of electromagnetic- and light-emission pulses in loaded materials.     

Diffusion in an ensemble of intersecting grain boundaries forming a triple junction

Grain boundary diffusion in an ensemble of three intersecting grain boundaries forming a triple junction is described in the framework of quasi-steady Fisher’s model. Two configurations, which differ in the number of grain boundaries adjacent to the surface with a diffusant source and in the tilt angle to the surface, are considered. Analytical expressions for the diffusant concentration distribution along each grain boundary that constitutes the triple junction and for the point of the triple junction are derived with the proviso of equal diffusion fluxes at the triple point. The expressions for the diffusant concentration distribution along the grain boundaries include not only diffusion constants (grain-boundary and bulk diffusion coefficients) but also structural characteristics of the ensemble of grain boundaries (the depth of the triple junction point and the angle between the grains forming the triple junction). It is shown that, if the coefficients of grain boundary diffusion are equal for all boundaries making the ensemble and for an equilibrium angle of 120° in a polycrystal, the diffusive mass transport rate in the triple junction zone is lower than that in a single grain boundary irrespective of the configuration of grain boundaries.     

Time-Expanded Sampling for Ensemble-Based Filters: Assimilation Experiments with Real Radar Observations

By sampling perturbed state vectors from each ensemble prediction run at properly selected time levels in the vicinity of the analysis time,the recently proposed time-expanded sampling approach can enlarge the ensemble size without increasing the number of prediction runs and,hence,can reduce the computational cost of an ensemble-based filter.In this study,this approach is tested for the first time with real radar data from a tornadic thunderstorm.In particular,four assimilation experiments were performed to test the time-expanded sampling method against the conventional ensemble sampling method used by ensemblebased filters.In these experiments,the ensemble square-root filter (EnSRF) was used with 45 ensemble members generated by the time-expanded sampling and conventional sampling from 15 and 45 prediction runs,respectively,and quality-controlled radar data were compressed into super-observations with properly reduced spatial resolutions to improve the EnSRF performances.The results show that the time-expanded sampling approach not only can reduce the computational cost but also can improve the accuracy of the analysis,especially when the ensemble size is severely limited due to computational constraints for real-radar data assimilation.These potential merits are consistent with those previously demonstrated by assimilation experiments with simulated data.     

Memory for multiple visual ensembles in infancy.

The number of individual items that can be maintained in working memory is limited. One solution to this problem is to store representations of ensembles that contain summary information about large numbers of items (e.g., the approximate number or cumulative area of a group of many items). Here we explored the developmental origins of ensemble representations by asking whether infants represent ensembles and, if so, how many at one time. We habituated 9-month-old infants to arrays containing 2, 3, or 4 spatially intermixed colored subsets of dots, then asked whether they detected a numerical change to one of the subsets or to the superset of all dots. Experiment Series 1 showed that infants detected a numerical change to 1 of the subsets when the array contained 2 subsets but not 3 or 4 subsets. Experiment Series 2 showed that infants detected a change to the superset of all dots no matter how many subsets were presented. Experiment 3 showed that infants represented both the approximate number and the cumulative surface area of these ensembles. Our results suggest that infants, like adults (Halberda, Sires, & Feigenson, 2006), can store quantitative information about 2 subsets plus the superset: a total of 3 ensembles. This converges with the known limit on the number of individual objects infants and adults can store and suggests that, throughout development, an ensemble functions much like an individual object for working memory. (PsycINFO Database Record (c) 2011 APA, all rights reserved)     

Frequency Analysis of a Linear Elastic Structure Carrying a Chain of Oscillators

In this technical note we analyze the free vibration of M undamped oscillators attached to an arbitrarily supported, linear elastic structure. Using the assumed-modes method with N component modes, the frequency equation governing the free vibration for this combined system is typically obtained as the characteristic determinant of a generalized eigenvalue problem of size (N + M) × (N + M). In this note we will show that by algebraically manipulating the generalized eigenvalue problem associated with free vibration, we can reduce it to a simple secular equation consisting of the sum of N terms, the roots or natural frequencies of which can be obtained either numerically or graphically. In addition, the resultant secular equation lends itself to the solution of an inverse problem that cannot be easily solved by analyzing the original generalized eigenvalue problem.     

Evaluating the energetics of empty cavities and internal mutations in proteins

The energetics of cavity formation in proteins is evaluated with two different approaches and results are analyzed and compared to experimental data. In the first approach, free energy of cavity formation is extracted by RMS fitting from the distribution of numbers of cavities, N, with different volumes, Vcav, in 80 high-resolution protein structures. It is assumed that the distribution of number of cavities according to their volume follows the Boltzmann law, N(Vcav) = exp [(-a.Vcav-b)/kT], or its simplified form. Specific energy cost of cavity formation, a, extracted by RMS fitting from these distributions is compared to a values extracted from experimental free energies of cavity formation in T4 lysozyme fitted to similar expressions. It is found that fitting of both sets of data leads to similar magnitudes and uncertainties in the calculated free energy values. It is shown that Boltzmann-like distribution of cavities can be derived for a simple model of an equilibrium interconversion between mutants in an extracellular system. We, however, suggest that a partitioning into cavity-dependent and cavity-independent terms may lose meaning when one attempts to describe mutation effects on protein stability in terms of specific free energy contributions. As an alternative approach, a direct molecular mechanics evaluation is attempted of T4 lysozyme destabilization by five single cavity-creating mutations. The calculations are based on the approach used in calculations of the energetics of packing defects in crystals. For all mutations calculated destabilizations agree with the corresponding experimental values within +/-0.6 kcal/mol. A computational relaxation of the mutant was most difficult to achieve for the mutation producing the smallest cavity. However, calculations do not always reproduce crystallographically observed contraction/expansion of cavities. It is suggested that this may be related to usually observed large RMS differences (> 1 A) between crystallographic and energy-minimized protein structures, and thus correct energetics might be easier to calculate than the correct geometry.     

Conditions for the appearance of a stable microcrack in the elastic field of a screened disclination

The conditions of possible microcrack nucleation and stability in the elastic field of disclinations screened by a free surface, disclinations of the opposite sense, or a distributed ensemble of moving dislocations are considered. The plastic deformation screening the elastic field of a disclination is shown to effectively stabilize microcrack growth at a sufficiently large grain size.     

Free energy formulation of fatigue crack initiation along persistent slip bands: calculation of SN curves and crack depths

A theory of fatigue crack initiation along PSBs based on free energy considerations has been developed. The PSB is modelled as an accumulation of vacancy dipoles. The total free energy of the system increases with fatigue cycle number due to the increase in elastic strain energy. It is shown that there exists a critical fatigue cycle number above which the initial state of dislocation accumulation along the slip band becomes energetically unstable, leading to the initiation of a microcrack within the PSB. SN curves for initiation can be calculated and the resulting crack depths determined. The theory correctly predicts several observed phenomena of fatigue crack initiation: (i) crack initiation is easier in air than in vacuum; (ii) crack initiation is easier at room temperature than at cryogenic temperatures; (iii) just-initiated cracks are shallow.     

Solution structure and backbone dynamics of component IV Glycera dibranchiata monomeric hemoglobin-CO

The solution structure and backbone dynamics of the recombinant, ferrous CO-ligated form of component IV monomeric hemoglobin from Glycera dibranchiata (GMH4CO) have been characterized by NMR spectroscopy. Distance geometry and simulated annealing calculations utilizing a total of 2550 distance and torsion angle constraints yielded an ensemble of 29 structures with an overall average backbone rmsd of 0.48 A from the average structure. Differences between the solution structure and a related crystal structure are confined to regions of lower precision in either the NMR or X-ray structure, or in regions where the amino acid sequences differ. 15N relaxation measurements at 76.0 and 60.8 MHz were analyzed with an extended model-free approach, and revealed low-frequency motions in the vicinity of the heme, concentrated in the F helix. Amide proton protection factors were obtained from H-D amide exchange measurements on 15N-labeled protein. Patterns in the backbone dynamics and protection factors were shown to correlate with regions of heterogeneity and disorder in the ensemble of NMR structures and with large crystallographic B-factors in the X-ray structures. Surprisingly, while the backbone atoms of the F helix have higher rmsds and larger measures of dynamics on the microsecond to millisecond time scale than the other helices, amide protection factors for residues in the F helix were observed to be similar to those of the other helices. This contrasts with H-D amide exchange measurements on sperm whale myoglobin which indicated low protection for the F helix (S. N. Loh and B. F. Volkman, unpublished results). These results for GMH4 suggest a model in which the F helix undergoes collective motions as a relatively rigid hydrogen-bonded unit, possibly pivoting about a central position near residue Val87.     

Perturbations of the denatured state ensemble: modeling their effects on protein stability and folding kinetics

By considering the denatured state of a protein as an ensemble of conformations with varying numbers of sequence-specific interactions, the effects on stability, folding kinetics, and aggregation of perturbing these interactions can be predicted from changes in the molecular partition function. From general considerations, the following conclusions are drawn: (1) A perturbation that enhances a native interaction in denatured state conformations always increases the stability of the native state. (2) A perturbation that promotes a non-native interaction in the denatured state always decreases the stability of the native state. (3) A change in the denatured state ensemble can alter the kinetics of aggregation and folding. (4) The loss (or increase) in stability accompanying two mutations, each of which lowers (or raises) the free energy of the denatured state, will be less than the sum of the effects of the single mutations, except in cases where both mutations affect the same set of partially folded conformations. By modeling the denatured state as the ensemble of all non-native conformations of hydrophobic-polar (HP) chains configured on a square lattice, it can be shown that the stabilization obtained from enhancement of native interactions derives in large measure from the avoidance of non-native interactions in the D state. In addition, the kinetic effects of fixing single native contacts in the denatured state or imposing linear gradients in the HH contact probabilities are found, for some sequences, to significantly enhance the efficiency of folding by a simple hydrophobic zippering algorithm. Again, the dominant mechanism appears to be avoidance of non-native interactions. These results suggest stabilization of native interactions and imposition of gradients in the stability of local structure are two plausible mechanisms involving the denatured state that could play a role in the evolution of protein folding and stability.     

Error estimates for results of nonstationary noise analysis derived with linear least squares methods

Nonstationary noise analysis of electrophysiological data is applied to the estimation of the single-channel current, i, and the number of active channels, N(C), whenever they cannot be determined directly due to limited resolution. Using least squares methods, the accuracy of estimating i and N(C) chiefly depends on the statistical error of the ensemble variance. It is shown that if the correlation among the binned data points is taken into account correctly, the variability of i and N(C) can be remarkably reduced and exact confidence limits of the parameters can be calculated. Least-squares methods are introduced which consider the measured error-covariance matrix of the binned variance in a model-independent fashion. Employing Monte Carlo methods, it is demonstrated that both the error predictions and the confidence limits are correct. The method is used to investigate the performance of nonstationary noise analysis at low channel open-probabilities. The application of the approach to simulated data as well as to experimental, i.e. non-ideal, data is discussed.     

Cross-linking studies and membrane localization and assembly of radiolabelled large mechanosensitive ion channel (MscL) of Escherichia coli

While the overall energy landscape of a foldable protein can be described by means of a few parameters characterizing its statistical topography, specific energetic terms subtly bias the representative structures giving rise to residue pair correlations as in a liquid. We use a free energy functional incorporating an inhomogeneous pair contact energy along with a contact formation entropy and a cooperativity contribution to determine residue-specific contact probabilities in the denatured state and the transition state ensemble. The predicted "hot residues" for the theoretical transition state ensemble reasonably agree with experiment for chymotrypsin inhibitor 2, and generally a strong correlation exists with the measured kinetic effects of mutating residues not involved in highly solvent-exposed regions.     

Substrate specificity of deubiquitinating enzymes: ubiquitin C-terminal hydrolases

Ubiquitin C-terminal hydrolases (UCH) are deubiquitinating enzymes which hydrolyze C-terminal esters and amides of ubiquitin. Here we report the processing of a number of ubiquitin derivatives by two human UCH isozymes (isozymes L1 and L3) and find that these enzymes show little discrimination based on the P1' amino acid, except that proline is cleaved slowly. Ubiquitinyllysine derivatives linked by the alpha- or epsilon-amino group are hydrolyzed at identical rates. Isozyme-specific hydrolytic preferences are only evident when the leaving group is large. The ubiquitin gene products can be cotranslationally processed by one or both of these UCH isozymes, and purified UbCEP52 can be hydrolyzed by UCH isozyme L3. Binding of nucleic acid by UbCEP52 converts it to a form resistant to processing by these enzymes, apparently because of the formation of a larger, more tightly folded substrate. Consistent with this postulate is the observation that these enzymes do not hydrolyze large ubiquitin derivatives such as N epsilon-ubiquitinyl-cytochrome-c, N epsilon-K48polyubiquitinyl-lysozyme, or an N alpha-ubiquitinyl-beta-galactosidase fusion protein. Thus, these enzymes rapidly and preferentially cleave small leaving groups such as amino acids and oligopeptides from the C-terminus of ubiquitin, but not larger leaving groups such as proteins. These data suggest that the physiological role of UCH is to hydrolyze small adducts of ubiquitin and to generate free monomeric ubiquitin from ubiquitin proproteins, but not to deubiquitinate ubiquitin-protein conjugates or disassemble polyubiquitin chains.     

The elastic rod model for DNA and its application to the tertiary structure of DNA minicircles in mononucleosomes

Explicit solutions to the equations of equilibrium in the theory of the elastic rod model for DNA are employed to develop a procedure for finding the configuration that minimizes the elastic energy of a minicircle in a mononucleosome with specified values of the minicircle size N in base pairs, the extent w of wrapping of DNA about the histone core particle, the helical repeat h(0)b of the bound DNA, and the linking number Lk of the minicircle. The procedure permits a determination of the set Y(N, w, h(0)b) of integral values of Lk for which the minimum energy configuration does not involve self-contact, and graphs of writhe versus w are presented for such values of Lk. For the range of N of interest here, 330 < N < 370, the set Y(N, w, h(0)b) is of primary importance: when Lk is not in Y(N, w, h(0)b), the configurations compatible with Lk have elastic energies high enough to preclude the occurrence of an observable concentration of topoisomer Lk in an equilibrium distribution of topoisomers. Equilibrium distributions of Lk, calculated by setting differences in the free energy of the extranucleosomal loop equal to differences in equilibrium elastic energy, are found to be very close to Gaussian when computed under the assumption that w is fixed, but far from Gaussian when it is assumed that w fluctuates between two values. The theoretical results given suggest a method by which one may calculate DNA-histone binding energies from measured equilibrium distributions of Lk.     

Forcing thermodynamically unfolded proteins to fold

A growing number of biologically important proteins have been identified as fully unfolded or partially disordered. Thus, an intriguing question is whether such proteins can be forced to fold by adding solutes found in the cells of some organisms. Nature has not ignored the powerful effect that the solution can have on protein stability and has developed the strategy of using specific solutes (called organic osmolytes) to maintain the structure and function cellular proteins in organisms exposed to denaturing environmental stresses (Yancey, P. H., Clark, M. E., Hand, S. C., Bowlus, R. D., and Somero, G. N. (1982) Science 217, 1214-1222). Here, we illustrate the extraordinary capability of one such osmolyte, trimethylamine N-oxide (TMAO), to force two thermodynamically unfolded proteins to fold to native-like species having significant functional activity. In one of these examples, TMAO is shown to increase the population of native state relative to the denatured ensemble by nearly five orders of magnitude. The ability of TMAO to force thermodynamically unstable proteins to fold presents an opportunity for structure determination and functional studies of an important emerging class of proteins that have little or no structure without the presence of TMAO.     

基于聚类欠采样的集成不均衡数据分类算法

传统的分类算法大多假设数据集是均衡的,追求整体的分类精度.而实际数据集经常是不均衡的,因此传统的分类算法在处理实际数据集时容易导致少数类样本有较高的分类错误率.现有针对不均衡数据集改进的分类方法主要有两类:一类是进行数据层面的改进,用过采样或欠采样的方法增加少数类数据或减少多数类数据;另一个是进行算法层面的改进.本文在原有的基于聚类的欠采样方法和集成学习方法的基础上,采用两种方法相结合的思想,对不均衡数据进行分类.即先在数据处理阶段采用基于聚类的欠采样方法形成均衡数据集,然后用AdaBoost集成算法对新的数据集进行分类训练,并在算法集成过程中引用权重来区分少数类数据和多数类数据对计算集成学习错误率的贡献,进而使算法更关注少数数据类,提高少数类数据的分类精度.     

Regularities in thermodynamic properties of interoxide polymeric compounds

Abstract

It is shown that in terms of the number N of elements in solid interoxide polymeric compounds and minerals, the molar heat capacity ratio C _/N is a _p single function of temperature. The molar heat capacity of a solid polymeric compound is found to be equal to the sum of the molar heat capacities of its constituent solid oxides at all temperatures. The standard entropies of formation of the alkali and alkaline earth polymeric compounds per atom of oxygen are both-95±5 J [atom O]- 1 K-1, which is similar to all other inorganic compounds. The standard free energies and enthalpies of formation of polymeric interoxide compounds at a given temperature T are directly proportional to the number N of elements in the compound.     

N-Demethylation reactions catalyzed by chloroperoxidase

The peroxidase-supported N-demethylations catalyzed by chloroperoxidase, a heme protein isolated from Caldariomyces fumago, have been investigated as models for cytochrome P-450-catalyzed N-dealkylations. The turnover number for the ethyl hydrogen peroxide-supported dealkylation of N,N-dimethylaniline by chloroperoxidase (1476) was much greater than that for cytochrome P-450-catalyzed dealkylations. The dealkylations of N,N-dimethylaniline by chloroperoxidase yielded N-methylaniline and formaldehyde in equimolar amounts with no other products detectable by high pressure liquid chromatography analysis of the reaction mixture. Ethyl hydrogen peroxidase could be replaced by other hydroperoxides, peroxides, or peracids. Chloride ions stimulated the reaction at low pH. The dealkylation reaction exhibited normal Michaelis-Menten saturation kinetics with respect to N,N-dimethylaniline (Km = 0.08 mM) and ethyl hydrogen peroxide (Km = 0.8 mM) at low substrate concentrations. However, substrate inhibition occurred at higher concentrations of N,N-dimethylaniline. The chloroperoxidase-catalyzed demethylations were inhibited by inhibitors of cytochrome P-450 such as azide or n-propyl gallate, but not by metyrapone, SKF-525A, or piperonyl butoxide. Although tiron and DL-epinephrine, trapping agents for the superoxide anion, inhibited the demethylation reactions, superoxide dismutase had no effect. There was no significant inhibition by alpha-phenyl-t-butyl-nitrone or 5,5-dimethyl-pyrroline-N-oxide, which react with free radicals. Diphenylfuran and DL-histidine, which react with singlet oxygen, did not inhibit the reaction. Substitution of D2O for H2O resulted in a marked inhibition with a solvent isotope effect (VH/VD) of 3.6. Chloroperoxidase did not catalyze the demethylation of N,N-dimethylaniline-N-oxide, indicating that the reaction does not proceed via an N-oxide intermediate.     

Influence of titanium and carbon contents on the hydrogen trapping of microalloyed steels

The presence of traps for hydrogen atoms influences the diffusivity and solubility of hydrogen itself in steels. In the present work, permeation measurements were carried out on hot-rolled microalloyed steels with different C and Ti contents in order to evaluate the number of irreversible and reversible traps. The number per unit volume of irreversible traps was correlated to calculated volume fraction of Ti(C,N) precipitates. These results, combined with microstructural investigations by transmission electron microscopy (TEM), showed that the largest number of irreversible traps was associated with steels having the largest volume fraction of fine and coherent Ti(C,N) precipitates. The reversible traps were associated with free Ti atoms, dislocations, and ferritic grain boundaries. Theoretical calculations confirmed the hydrogen binding energy of Ti free atoms (−27.1 kJ/mol).