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Long-range HNCO NMR spectra for proteins show crosspeaks due to 1JNC', 2JNC', 3JNCγ, and h3JNC' couplings. The h3JNC' couplings are transmitted through hydrogen bonds and their sizes are correlated to hydrogen bond lengths. We collected long-range HNCO data at a series of temperatures for four protein structures. P22i and CUS-3i are six-stranded beta-barrel I-domains from phages P22 and CUS-3 that share less than 40% sequence identity. The cis and trans states of the C-terminal domain from pore-forming toxin hemolysin ΙΙ (HlyIIC) arise from the isomerization of a single G404-P405 peptide bond. For P22i and CUS-3i, hydrogen bonds detected by NMR agree with those observed in the corresponding domains from cryoEM structures of the two phages. Hydrogen bond lengths derived from the h3JNC' couplings, however, are poorly conserved between the distantly related CUS-3i and P22i domains and show differences even between the closely related cis and trans state structures of HlyIIC. This is consistent with hydrogen bond lengths being determined by local differences in structure rather than the overall folding topology. With increasing temperature, hydrogen bonds typically show an apparent increase in length that has been attributed to protein thermal expansion. Some hydrogen bonds are invariant with temperature, however, while others show apparent decreases in length, suggesting they become stabilized with increasing temperature. Considering the data for the three proteins in this study and previously published data for ubiquitin and GB3, lowered protein folding stability and cooperativity corresponds with a larger range of temperature responses for hydrogen bonds. This suggests a partial uncoupling of hydrogen bond energetics from global unfolding cooperativity as protein stability decreases. [ABSTRACT FROM AUTHOR]

Long-range HNCO NMR spectra for proteins show crosspeaks due to 1JNC′, 2JNC′, 3JNCγ, and h3JNC′ couplings. The h3JNC′ couplings are transmitted through hydrogen bonds and their sizes are correlated to hydrogen bond lengths. We collected long-range HNCO data at a series of temperatures for four protein structures. P22i and CUS-3i are six-stranded beta-barrel I-domains from phages P22 and CUS-3 that share less than 40% sequence identity. The cis and trans states of the C-terminal domain from pore-forming toxin hemolysin ΙΙ (HlyIIC) arise from the isomerization of a single G404-P405 peptide bond. For P22i and CUS-3i, hydrogen bonds detected by NMR agree with those observed in the corresponding domains from cryoEM structures of the two phages. Hydrogen bond lengths derived from the h3JNC′ couplings, however, are poorly conserved between the distantly related CUS-3i and P22i domains and show differences even between the closely related cis and trans state structures of HlyIIC. This is consistent with hydrogen bond lengths being determined by local differences in structure rather than the overall folding topology. With increasing temperature, hydrogen bonds typically show an apparent increase in length that has been attributed to protein thermal expansion. Some hydrogen bonds are invariant with temperature, however, while others show apparent decreases in length, suggesting they become stabilized with increasing temperature. Considering the data for the three proteins in this study and previously published data for ubiquitin and GB3, lowered protein folding stability and cooperativity corresponds with a larger range of temperature responses for hydrogen bonds. This suggests a partial uncoupling of hydrogen bond energetics from global unfolding cooperativity as protein stability decreases.

While it is widely appreciated that the denatured state of a protein is a heterogeneous conformational ensemble, there is still debate over how this ensemble changes with environmental conditions. Here, we use single-molecule chemo-mechanical unfolding, which combines force and urea using the optical tweezers, together with traditional protein unfolding studies to explore how perturbants commonly used to unfold proteins (urea, force, and temperature) affect the denatured-state ensemble. We compare the urea m-values, which report on the change in solvent accessible surface area for unfolding, to probe the denatured state as a function of force, temperature, and urea. We find that while the urea- and force-induced denatured states expose similar amounts of surface area, the denatured state at high temperature and low urea concentration is more compact. To disentangle these two effects, we use destabilizing mutations that shift the Tm and Cm. We find that the compaction of the denatured state is related to changing temperature as the different variants of acyl-coenzyme A binding protein have similar m-values when they are at the same temperature but different urea concentration. These results have important implications for protein folding and stability under different environmental conditions.

Abstract Background High-throughput technologies enable the cost-effective collection and analysis of DNA methylation data throughout the human genome. This naturally entails missing values management that can complicate the analysis of the data. Several general and specific imputation methods are suitable for DNA methylation data. However, there are no detailed studies of their performances under different missing data mechanisms –(completely) at random or not- and different representations of DNA methylation levels (β and M-value). Results We make an extensive analysis of the imputation performances of seven imputation methods on simulated missing completely at random (MCAR), missing at random (MAR) and missing not at random (MNAR) methylation data. We further consider imputation performances on the popular β- and M-value representations of methylation levels. Overall, β-values enable better imputation performances than M-values. Imputation accuracy is lower for mid-range β-values, while it is generally more accurate for values at the extremes of the β-value range. The MAR values distribution is on the average more dense in the mid-range in comparison to the expected β-value distribution. As a consequence, MAR values are on average harder to impute. Conclusions The results of the analysis provide guidelines for the most suitable imputation approaches for DNA methylation data under different representations of DNA methylation levels and different missing data mechanisms.

An asphalt-binder is one of the key constituent elements that significantly influences and controls the fracture behavior and cracking performance of hot-mix asphalt (HMA). In particular, the low-temperature rheological properties of asphalt-binders are critical in terms of improving the HMA fracture properties in the laboratory and ultimately mitigating cracking in the field. The bending beam rheometer (BBR) is one of the most widely used laboratory test methods for evaluating and quantifying the asphalt-binder behavior at low temperatures, mostly using the stiffness (S) and m-value parameters. Using the Texas flexible pavements and overlays database, namely, the Texas data storage system (DSS), as the primary data source, this study was conducted to correlate the asphalt-binder low-temperature rheological properties (measured using the BBR) to the HMA fracture properties. The HMA fracture properties were measured in the laboratory using the monotonic-loading overlay tester (ML-OT). In general, the asphalt-binder low-temperature rheological properties [particularly the stiffness (S)] exhibited promising potential to predict the HMA fracture properties, namely, the tensile strain (εt) and fracture energy index (FE Index), with a coefficient of determination (R2) greater than 60%. [ABSTRACT FROM AUTHOR]

Long-range HNCO NMR spectra for proteins show crosspeaks due to 1 J NC' , 2 J NC' , 3 J NCγ , and h3 J NC' couplings. The h3 J NC' couplings are transmitted through hydrogen bonds and their sizes are correlated to hydrogen bond lengths. We collected long-range HNCO data at a series of temperatures for four protein structures. P22i and CUS-3i are six-stranded beta-barrel I-domains from phages P22 and CUS-3 that share less than 40% sequence identity. The cis and trans states of the C-terminal domain from pore-forming toxin hemolysin ΙΙ (HlyIIC) arise from the isomerization of a single G404-P405 peptide bond. For P22i and CUS-3i, hydrogen bonds detected by NMR agree with those observed in the corresponding domains from cryoEM structures of the two phages. Hydrogen bond lengths derived from the h3 J NC' couplings, however, are poorly conserved between the distantly related CUS-3i and P22i domains and show differences even between the closely related cis and trans state structures of HlyIIC. This is consistent with hydrogen bond lengths being determined by local differences in structure rather than the overall folding topology. With increasing temperature, hydrogen bonds typically show an apparent increase in length that has been attributed to protein thermal expansion. Some hydrogen bonds are invariant with temperature, however, while others show apparent decreases in length, suggesting they become stabilized with increasing temperature. Considering the data for the three proteins in this study and previously published data for ubiquitin and GB3, lowered protein folding stability and cooperativity corresponds with a larger range of temperature responses for hydrogen bonds. This suggests a partial uncoupling of hydrogen bond energetics from global unfolding cooperativity as protein stability decreases.

The Illumina Infinium HumanMethylation450 Beadchips have been widely utilized in epigenome-wide association studies (EWAS). However, the existing two types of probes (type I and type II), with the distribution of measurements of probes and dynamic range different, may bias downstream analyses. Here, we propose a method, MGMIN (M-values Gaussian-MIxture Normalization), to correct the probe designs based on M-values of DNA methylation. Our strategy includes fitting Gaussian mixture distributions to type I and type II probes separately, a transformation of M-values into quantiles and finally a dilation transformation based on M-values of DNA methylation to maintain the continuity of the data. Our method is validated on several public datasets on reducing probe design bias, reducing the technical variation and improving the ability to find biologically differential methylation signals. The results show that MGMIN achieves competitive performances compared to BMIQ which is a well-known normalization method on β-values of DNA methylation.

The Illumina Infinium HumanMethylation450 Beadchips have been widely utilized in epigenome-wide association studies (EWAS). However, the existing two types of probes (type I and type II), with the distribution of measurements of probes and dynamic range different, may bias downstream analyses. Here, we propose a method, MGMIN (M -values Gaussian-MIxture Normalization), to correct the probe designs based on M -values of DNA methylation. Our strategy includes fitting Gaussian mixture distributions to type I and type II probes separately, a transformation of M -values into quantiles and finally a dilation transformation based on M -values of DNA methylation to maintain the continuity of the data. Our method is validated on several public datasets on reducing probe design bias, reducing the technical variation and improving the ability to find biologically differential methylation signals. The results show that MGMIN achieves competitive performances compared to BMIQ which is a well-known normalization method on β-values of DNA methylation. [ABSTRACT FROM AUTHOR]

Background: High-throughput technologies enable the cost-effective collection and analysis of DNA methylation data throughout the human genome. This naturally entails missing values management that can complicate the analysis of the data. Several general and specific imputation methods are suitable for DNA methylation data. However, there are no detailed studies of their performances under different missing data mechanisms –(completely) at random or not- and different representations of DNA methylation levels (β and M-value). Results: We make an extensive analysis of the imputation performances of seven imputation methods on simulated missing completely at random (MCAR), missing at random (MAR) and missing not at random (MNAR) methylation data. We further consider imputation performances on the popular β- and M-value representations of methylation levels. Overall, β-values enable better imputation performances than M-values. Imputation accuracy is lower for mid-range β-values, while it is generally more accurate for values at the extremes of the β-value range. The MAR values distribution is on the average more dense in the mid-range in comparison to the expected β-value distribution. As a consequence, MAR values are on average harder to impute. Conclusions: The results of the analysis provide guidelines for the most suitable imputation approaches for DNA methylation data under different representations of DNA methylation levels and different missing data mechanisms. [ABSTRACT FROM AUTHOR]

The present article investigates in laboratory conditions the effect of cyclic freezing and thawing (CFT) on characteristics of the neat bitumen and bitumens modified with different polymers, as well as stone mastic asphalt concretes prepared with the use of the neat and the modified bitumens. Maximal and minimal temperature in cycles of freezing and thawing (FT) is accepted equal to +20 °C and −18 ± 2 °C respectively. Characteristics of low temperature resistance of bitumens (stiffness S and m-value) have been determined on bending beam rheometer at the temperatures of −24 °C, −30 °C, and −36 °C. Strength at direct tension of the stone mastic asphalt concretes has been evaluated in the device TRAVIS (Infratest Ltd., Brackenheim, Germany) at the temperatures of 0 °C, −10 °C, −20 °C, and −30 °C. Resistance of stone mastic asphalt concretes to rutting has been determined on a Hamburg wheel tracking machine. The characteristics of the bitumens have been determined in the initial condition and after 25 and 50 cycles of FT, and the characteristics of stone mastic asphalt concretes, in the initial condition and after 50 cycles of FT. The results obtained experimentally have shown that CFT impacts essentially on the investigated characteristics of bitumens and stone mastic asphalt concretes. Modification with polymers improves the mechanical characteristics of the bitumens and decreases the effect of CFT. It is also found out that when selecting a bitumen for specific climatic conditions it is necessary to take into account both the number of cycles of FT and the characteristic low temperature. [ABSTRACT FROM AUTHOR]

The present article investigates in laboratory conditions the effect of cyclic freezing and thawing (CFT) on characteristics of the neat bitumen and bitumens modified with different polymers, as well as stone mastic asphalt concretes prepared with the use of the neat and the modified bitumens. Maximal and minimal temperature in cycles of freezing and thawing (FT) is accepted equal to +20 °C and −18 ± 2 °C respectively. Characteristics of low temperature resistance of bitumens (stiffness S and m-value) have been determined on bending beam rheometer at the temperatures of −24 °C, −30 °C, and −36 °C. Strength at direct tension of the stone mastic asphalt concretes has been evaluated in the device TRAVIS (Infratest Ltd., Brackenheim, Germany) at the temperatures of 0 °C, −10 °C, −20 °C, and −30 °C. Resistance of stone mastic asphalt concretes to rutting has been determined on a Hamburg wheel tracking machine. The characteristics of the bitumens have been determined in the initial condition and after 25 and 50 cycles of FT, and the characteristics of stone mastic asphalt concretes, in the initial condition and after 50 cycles of FT. The results obtained experimentally have shown that CFT impacts essentially on the investigated characteristics of bitumens and stone mastic asphalt concretes. Modification with polymers improves the mechanical characteristics of the bitumens and decreases the effect of CFT. It is also found out that when selecting a bitumen for specific climatic conditions it is necessary to take into account both the number of cycles of FT and the characteristic low temperature.

Reclaimed asphalt pavement (RAP) has many advantages and is utilized to improve the high temperature properties of asphalt mixtures. Low temperature cracking is a predominant distress in asphalt pavements containing RAP materials. Thus, the evaluation of fracture resistance for asphalt mixtures containing RAP is of interest. The objective of this research is to explore the low temperature performance characteristics of RAP mortars containing sieved RAP and soft binders at three aged states. The stiffness values and m-values from bending beam rheometer (BBR) tests at three test temperatures of -18 °C, -12 °C and -6 °C were obtained to conduct the minimum low temperature grades. RAP mortar with a higher aged binder content had a higher minimum low temperature regardless of RAP source. In addition, RAP mortars with virgin soft binder had the best low temperature resistance followed by the RAP mortars with rolling thin film oven (RTFO) and pressure-aged vessel (PAV) binders. [ABSTRACT FROM AUTHOR]

Small solutes affect protein and nucleic acid processes because of favorable or unfavorable chemical interactions of the solute with the biopolymer surface exposed or buried in the process. Large solutes also exclude volume and affect processes where biopolymer molecularity and/or shape changes. Here, we develop an analysis to separate and interpret or predict excluded volume and chemical effects of a flexible coil polymer on a process. We report a study of the concentration-dependent effects of the full series from monomeric to polymeric PEG on intramolecular hairpin and intermolecular duplex formation by 12-nucleotide DNA strands. We find that chemical effects of PEG on these processes increase in proportion to the product of the amount of DNA surface exposed on melting and the amount of PEG surface that is accessible to this DNA, and these effects are completely described by two interaction terms that quantify the interactions between this DNA surface and PEG end and interior groups. We find that excluded volume effects, once separated from these chemical effects, are quantitatively described by the analytical theory of Hermans, which predicts the excluded volume between a flexible polymer and a rigid molecule. From this analysis, we show that at constant concentration of PEG monomer, increasing PEG size increases the excluded volume effect but decreases the chemical interaction effect, because in a large PEG coil a smaller fraction of the monomers are accessible to the DNA. Volume exclusion by PEG has a much larger effect on intermolecular duplex formation than on intramolecular hairpin formation. [ABSTRACT FROM AUTHOR]

Abstract: Thermal unfolding of ribonuclease A and α-chymotrypsinogen A was analyzed in various alcohol solutions of methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, 2-butanol, tert-butanol, trifluoroethanol, and glycerol. The change in thermal unfolding ratio with temperature was described well by the van''t Hoff equation and the melting temperature and the enthalpy of protein unfolding were obtained. The reciprocal form of the Wyman–Tanford equation, which describes the unfolded-to-folded protein ratio as a function of water activity, was applied to obtain a linear plot. From the slope of this plot and water activity, the stabilization free energy (ΔΔG) in a solution was calculated. This shows an important role of water activity in protein stability. ΔΔG was linearly dependent on alcohol concentration and m-values of alcohols for protein unfolding were obtained. This provides a theoretical basis for the linear extrapolation model (LEM). The m-values for alcohols were negative except for glycerol. The negative higher m-value for longer and linear chain alcohols suggested the important role of the disturbance of hydrophobic interactions as well as the hydrogen-bonding in the mechanism of protein destabilization by alcohols. The number of change in bound-alcohol molecules upon protein unfolding was also obtained. [Copyright &y& Elsevier]

Analyzing the stability of a multimeric protein is challenging because of the intrinsic difficulty in handling the mathematical model for the folded multimer-unfolded monomer equilibrium. To circumvent this problem, we introduce the concept of effective stability, Δ Geff (= − RTln Keff), where Keff is the equilibrium constant expressed in monomer units. Analysis of the denaturant effect on Δ Geff gives new insight into the stability of multimeric proteins. When a multimeric protein is mostly folded, the dependence of effective stability on denaturant concentration (effective m-value) is simply the m-value of its monomeric unit. However, when the protein is mostly unfolded, its stability depends on denaturant concentration with the m-value of its multimeric form. We also find that the effective m-value at the C m is a good approximation of the apparent m-value determined by fitting the equilibrium unfolding data from multimeric proteins with a two-state monomer model. Moreover, when the m-value of a monomeric unit is estimated from its size, the effective stability of a multimeric protein can be determined simply from C m and this estimated m-value. These simple and intuitive approaches will allow a facile analysis of the stability of multimeric proteins. These analyses are also applicable for high-throughput analysis of protein stability on a proteomic scale. [ABSTRACT FROM AUTHOR]

Three hundred and twenty eight serial measurements of blood glucose (SMBG) performed in a Department of Medicine were rated by 7 experienced diabetes specialists. A statistical study showed considerable variation among the raters. Similarly, the ratings of the 7 physicians relative to the mean rating showed a great deal of individual variation. A highly satisfactory correlation was found between the mean rating of the 7 physicians and the M-value of Schlichtkrull as computed from 7 or 8 samples collected during the day. On the basis of these findings the degree of diabetes control for this population can be judged satisfactory, good or very good when the M-value is ⩽ 10. If the M-value is ⩾ 22, control is poor or very poor and major changes in the treatment program are indicated. Intermediate values, seen in one out of five cases, indicate the need for systematic consultation with a diabetes specialist. [ABSTRACT FROM AUTHOR]