Net Charge Values Research Articles

Hijacking of N-fixing legume albumin-1 genes enables the cyclization and stabilization of defense peptides

The legume albumin-1 gene family, arising after nodulation, encodes linear a- and b-chain peptides for nutrient storage and defense. Intriguingly, in one prominent legume, Clitoria ternatea, the b-chains are replaced by domains producing ultra-stable cyclic peptides called cyclotides. The mechanism of this gene hijacking is until now unknown. Cyclotides require recruitment of ligase-type asparaginyl endopeptidases (AEPs) for maturation (cyclization), necessitating co-evolution of two gene families. Here we compare a chromosome-level C. ternatea genome with grain legumes to reveal an 8 to 40-fold expansion of the albumin-1 gene family, enabling the additional loci to undergo diversification. Iterative rounds of albumin-1 duplication and diversification create four albumin-1 enriched genomic islands encoding cyclotides, where they are physically grouped by similar pI and net charge values. We identify an ancestral hydrolytic AEP that exhibits neofunctionalization and multiple duplication events to yield two ligase-type AEPs. We propose cyclotides arise by convergence in C. ternatea where their presence enhances defense from biotic attack, thus increasing fitness compared to lineages with linear b-chains and ultimately driving the replacement of b-chains with cyclotides.

Self-Interaction of Human Serum Albumin: A Formulation Perspective.

In the present study, small-angle X-ray scattering (SAXS) and static light scattering (SLS) have been used to study the solution properties and self-interaction of recombinant human serum albumin (rHSA) molecules in three pharmaceutically relevant buffer systems. Measurements are carried out up to high protein concentrations and as a function of ionic strength by adding sodium chloride to probe the role of electrostatic interactions. The effective structure factors (Seff) as a function of the scattering vector magnitude q have been extracted from the scattering profiles and fit to the solution of the Ornstein–Zernike equation using a screened Yukawa potential to describe the double-layer force. Although only a limited q range is used, accurate fits required including an electrostatic repulsion element in the model at low ionic strength, while only a hard sphere model with a tunable diameter is necessary for fitting to high-ionic-strength data. The fit values of net charge agree with available data from potentiometric titrations. Osmotic compressibility data obtained by extrapolating the SAXS profiles or directly from SLS measurements has been fit to a 10-term virial expansion for hard spheres and an equation of state for hard biaxial ellipsoids. We show that modeling rHSA as an ellipsoid, rather than a sphere, provides a much more accurate fit for the thermodynamic data over the entire concentration range. Osmotic virial coefficient data, derived at low protein concentration, can be used to parameterize the model for predicting the behavior up to concentrations as high as 450 g/L. The findings are especially important for the biopharmaceutical sector, which require approaches for predicting concentrated protein solution behavior using minimal sample consumption.

Effect of ultrasound pre-treatment on the characterization and properties of collagen extracted from soft-shelled turtle (Pelodiscus sinensis)

The aim of the present study was to evaluate the effect of ultrasound pre-treatment on the characterization and in-vitro digestion properties of collagen from turtle calipash and the antioxidant activity of its hydrolysates. The results showed that ultrasound significantly increased the collagen yield compared with that from conventional extraction (ASC, P < 0.05). Containing α1 and α2 chains, acid-soluble collagen (ASC) and the collagen with ultrasound pre-treatment (UASC) were characterized as type I collagen in SDS-PAGE patterns and UASC also maintained their structural integrity in the circular dichroism spectra. At the same pH, UASC had smaller particle size and larger absolute value of net charge than ASC. Moreover, ultrasound could induce apparent physical changes as looser, more porous and homogenous in microstructures imagines. Influenced by various pH, UASC showed higher solubility than ASC. Additionally, UASC was more prone to enzymolysis in vitro digestion and their hydrolysates showed significant increases antioxidant properties over ASC (P < 0.05), indicating that UASC would be a potential substrate to generate bioactive peptides. These results suggested that UASC would be a potential new material for biomedical applications and functional supplement in food industries.

Revisiting Benzylidenequinolinylnickel Catalysts through the Electronic Effects on Catalytic Activity by DFT Studies

The reaction activities for a series of benzylidenequinolin nickel complex systems are studied by density functional theory methods. The effective net charge values obtained on nickel atoms illustrate the correlation with experimental activities. Catalytic activity increases with the higher effective net charge values. The energy gaps (epsilon(1)) between Ni complex's lowest unoccupied molecular orbita (LUMO) and ethylene's highest occupied molecular orbital (HOMO) orbitals are calculated, regarding both pre-catalysts and active species. The results show that there are also relationships between epsilon(1) and catalytic activities. Comparing the pre-catalysts to the active species, the energy gaps epsilon(1) decrease dramatically from about 60 kcal mol(-1) to about 0 kcal mol(-1) for each complex, indicating the important activating role played by the co-catalyst. Finally, it is found that the energy difference values between different spin states relates to catalytic activities for both pre-catalyst and active species as well.

V3 net charge: additional tool in HIV-1 tropism prediction.

Genotype-based algorithms are valuable tools for the identification of patients eligible for CCR5 inhibitors administration in clinical practice. Among the available methods, geno2pheno[coreceptor] (G2P) is the most used online tool for tropism prediction. This study was conceived to assess if the combination of G2P prediction with V3 peptide net charge (NC) value could improve the accuracy of tropism prediction. A total of 172 V3 bulk sequences from 143 patients were analyzed by G2P and NC values. A phenotypic assay was performed by cloning the complete env gene and tropism determination was assessed on U87_CCR5(+)/CXCR4(+) cells. Sequences were stratified according to the agreement between NC values and G2P results. Of sequences predicted as X4 by G2P, 61% showed NC values higher than 5; similarly, 76% of sequences predicted as R5 by G2P had NC values below 4. Sequences with NC values between 4 and 5 were associated with different G2P predictions: 65% of samples were predicted as R5-tropic and 35% of sequences as X4-tropic. Sequences identified as X4 by NC value had at least one positive residue at positions known to be involved in tropism prediction and positive residues in position 32. These data supported the hypothesis that NC values between 4 and 5 could be associated with the presence of dual/mixed-tropic (DM) variants. The phenotypic assay performed on a subset of sequences confirmed the tropism prediction for concordant sequences and showed that NC values between 4 and 5 are associated with DM tropism. These results suggest that the combination of G2P and NC could increase the accuracy of tropism prediction. A more reliable identification of X4 variants would be useful for better selecting candidates for Maraviroc (MVC) administration, but also as a predictive marker in coreceptor switching, strongly associated with the phase of infection.

Correlating net charges and the activity of bis(imino)pyridylcobalt complexes in ethylene polymerization

A series of unsymmetrical bis(imino)pyridylcobalt complexes was investigated by DFT–QEq method to correlate the net charges of cobalt center and their catalytic activities. The effective net charge values on cobalt atom, calculated by QEq method at doublet spin state, reflected the influences of substituents within their ligands. The catalytic activities of these cobalt complexes in ethylene polymerization continuously increase along with higher values of the effective net charges on cobalt atom. The energy differences at doublet and quartet states kept the constant tendency for the cobalt complexes without the steric influences of substituents within ligands.

Protein charge ladders reveal that the net charge of ALS-linked superoxide dismutase can be different in sign and magnitude from predicted values.

This article utilized "protein charge ladders"-chemical derivatives of proteins with similar structure, but systematically altered net charge-to quantify how missense mutations that cause amyotrophic lateral sclerosis (ALS) affect the net negative charge (Z) of superoxide dismutase-1 (SOD1) as a function of subcellular pH and Zn(2+) stoichiometry. Capillary electrophoresis revealed that the net charge of ALS-variant SOD1 can be different in sign and in magnitude-by up to 7.4 units per dimer at lysosomal pH-than values predicted from standard pKa values of amino acids and formal oxidation states of metal ions. At pH 7.4, the G85R, D90A, and G93R substitutions diminished the net negative charge of dimeric SOD1 by up to +2.29 units more than predicted; E100K lowered net charge by less than predicted. The binding of a single Zn(2+) to mutant SOD1 lowered its net charge by an additional +2.33 ± 0.01 to +3.18 ± 0.02 units, however, each protein regulated net charge when binding a second, third, or fourth Zn(2+) (ΔZ < 0.44 ± 0.07 per additional Zn(2+) ). Both metalated and apo-SOD1 regulated net charge across subcellular pH, without inverting from negative to positive at the theoretical pI. Differential scanning calorimetry, hydrogen-deuterium exchange, and inductively coupled plasma mass spectrometry confirmed that the structure, stability, and metal content of mutant proteins were not significantly affected by lysine acetylation. Measured values of net charge should be used when correlating the biophysical properties of a specific ALS-variant SOD1 protein with its observed aggregation propensity or clinical phenotype.

Improving HIV‐1 tropism determination by combining geno2pheno and V3 net charge calculation

Genotypic tests are the most common methods to identify patients eligible for CCR5 inhibitors administration in Europe. Among the available tools geno2pheno coreceptor (G2P) is the most used online system in routine diagnostics. This study was conceived to assess if the combination of G2P prediction with V3 peptide net charge (NC) value could improve the accuracy of tropism prediction. Sequences (129) were analyzed by G2P according to European Guidelines. NC values were calculated by the online software Peptide Property Calculator. Phenotypic assay was performed cloning the complete env gene into pcDNA 3.1 TOPO vector; infectivity of pseudotyped virions was tested on U87_CD4+CCR5+ and U87_CD4+CXCR4+ cells lines to assess viral tropism. Sequences were stratified into 3 groups according to the agreement between NC values and G2P results. Group 1: sequences assigned to the same group by both tools, group 2: sequences assigned to one group by G2P but indeterminate by NC and group 3: sequences for which G2P and NC gave discordant results. 61% of sequences predicted as X4 by G2P showed NC values higher than 5; similarly, 76% of sequences predicted as R5 by G2P had NC values below 4 (Group 1). Sequences with NC values between 4 and 5 (Group 2) were associated to different G2P predictions: 59% samples were predicted as R5-tropic and 41% sequences as X4-tropic. These data support the hypothesis that 4 to 5 NC values could be associated to the presence of dual/mixed-tropic variants (DM). Sequences identified as X4 by NC value had at least one positive residue in positions known to be involved in tropism prediction (58%) and positive residues in position 32 (39%). To further verify NC-based prediction, phenotypic assay was performed on a subset of sequences from each group. The assay confirmed the tropism prediction for group 1 sequences and demonstrated that the variants with net charge between 4 and 5 have DM tropism. Moreover, in vitro phenotyping of discordant viruses confirmed NC result, showing that this parameter is strongly associated with phenotypic assay. These results show that the combination of G2P and NC could increase the accuracy of tropism prediction and the ability to discriminate DM viruses. A more reliable identification of X4 variants would be useful for better selecting candidates for maraviroc administration, but also as a predictive marker in coreceptor switching, strongly associated to the phase of infection.

On the Zwitterionic Nature of Gas-Phase Peptides and Protein Ions

Determining the total number of charged residues corresponding to a given value of net charge for peptides and proteins in gas phase is crucial for the interpretation of mass-spectrometry data, yet it is far from being understood. Here we show that a novel computational protocol based on force field and massive density functional calculations is able to reproduce the experimental facets of well investigated systems, such as angiotensin II, bradykinin, and tryptophan-cage. The protocol takes into account all of the possible protomers compatible with a given charge state. Our calculations predict that the low charge states are zwitterions, because the stabilization due to intramolecular hydrogen bonding and salt-bridges can compensate for the thermodynamic penalty deriving from deprotonation of acid residues. In contrast, high charge states may or may not be zwitterions because internal solvation might not compensate for the energy cost of charge separation.

New Model for Polymerization of Oligomeric Alcohol Dehydrogenases into Nanoaggregates

Polymerization and self-assembly of proteins into nanoaggregates of different sizes and morphologies (nanoensembles or nanofilaments) is a phenomenon that involved problems in various neurodegenerative diseases (medicine) and enzyme instability/inactivity (biotechnology). Thermal polymerization of horse liver alcohol dehydrogenase (dimeric) and yeast alcohol dehydrogenase (tetrameric), as biotechnological ADH representative enzymes, was evaluated for the development of a rational strategy to control aggregation. Constructed ADH nuclei, which grew to larger amorphous nanoaggregates, were prevented via high repulsion strain of the net charge values. Good correlation between the variation in scattering and lambda(-2) was related to the amorphousness of the nanoaggregated ADHs, shown by electron microscopic images. Scattering corrections revealed that ADH polymerization was related to the quaternary structural changes, including delocalization of subunits without unfolding, i.e. lacking the 3D conformational and/or secondary-ordered structural changes. The results demonstrated that electrostatic repulsion was not only responsible for disaggregation but also caused a delay in the onset of aggregation temperature, decreasing maximum values of aggregation and amounts of precipitation. Together, our results demonstrate and propose a new model of self-assembly for ADH enzymes based on the construction of nuclei, which grow to formless nanoaggregates with minimal changes in the tertiary and secondary conformations.

Molecular dynamics simulations of tubulin structure and calculations of electrostatic properties of microtubules

We present the results of molecular dynamics computations based on the atomic resolution structure of tubulin. Values of net charge, charge distribution and dipole moment components are obtained for the tubulin heterodimer. Physical consequences of these results are discussed for microtubules in terms of the effects on test charges, test dipoles, and neighboring microtubules.

Polyamine류와 Imidazole 유도체가 배위된 Ni(Ⅱ) 착물의 저해 효과에 관한 분자궤도함수론적 해석

폴리아민과 이미다졸 유도체에 대해서 AM1, PM3, ZINDO/1법으로 형성엔탈피(<TEX>${\Delta}H_f$</TEX>), HOMO, LUMO에너지 및 쌍극자힘(<TEX>${\mu}_D$</TEX>) 값을 얻었다. 이들 리간드를 Ni(II)에 배위시킨 Ni(II)착물의 저해활성효과를 ZINDO/1(Zerner''s Intermediate Neglect of Differential Overlap No.1)법으로 조사하였다. Ni(II)착물의 중심금속의 알짜전하 값이 0.03 이상과 형성엔탈피가 음의 값으로 300 eV이상에서 실험적인 저해활성 값이 나타남을 보였다. 그 결과 Ni(II)착물은 불안정한 5배위 사각피라미드형인 [Ni(dpt)(tn)]<TEX>$^{2+}$</TEX>(dpt=3,3''-diaminodipropylamine)(tn=1,3-diaminopropane)와 일그러진 사면체인 [Ni(N-PropIm)<TEX>$_2$</TEX>(NCS))<TEX>$_2$</TEX>](N-PropIm=N-Propylimidazole)를 제외한 착물에서는 저해활성 값이 나타남을 알 수 있었다. Quantum chemical quantities, enthalpy of formation(<TEX>${\Delta}H_f$</TEX>), HOMO and LUMO energy, and dipole moment(<TEX>${\mu}_D$</TEX>) were acquired by AM1, PM3, and ZINDO/1 methods for polyamines and imidazole derivatives. The investigation of the inhibitory activity on some Ni(II) complexes with polyamines and imidazole derivatives is performed by ZINDO/1 calculations. It was found that experimental inhibitory activity(IA) appeared when the value of net charge and enthalpy of formation were over 0.03 and -300 eV, respectively for Ni(II) complexes. These results showed that the Ni(II) complexes have exception on the following very unstable compounds: square pyramidal [Ni(dpt)(tn)])]<TEX>$^{2+}$</TEX>(dpt=3,3'-diaminodipropylamine)(tn=1,3-diaminopropane) and distorted tetrahedral [Ni(N-PropIm))<TEX>$_2$</TEX>(NCS))<TEX>$_2$</TEX>](N-PropIm=N-Propylimidazole).

Improvement of High-temperature Creep Resistance in Polycrystalline Al&lt;SUB&gt;2&lt;/SUB&gt;O&lt;SUB&gt;3&lt;/SUB&gt; by Cations Co-doping

High-temperature creep resistance in cations co-doped polycrystalline Al 2 O 3 was examined by uniaxial compression creep test at 1250°C. The dopant oxides used in this study were 0.1 mol% of YO 1 . 5 , ZrO 2 , SrO, MgO and TiO 2 . The creep rate in Al 2 O 3 was significantly changed by cations co-doping. For instance, Zr/Y co-doping suppressed the creep rate in Al 2 O 3 by a factor of about 400. A high-resolution transmission electron microscopy (HREM) and nano-probe energy dispersive X-ray spectroscopy (EDS) analysis revealed that Y and Zr cations segregate along grain boundaries. The grain boundary diffusion in Al 2 O 3 was supposed to be retarded by the segregation of Y and Zr cations. A first-principle molecular orbital calculation was made for cations co-doped Al 2 O 3 and cation singly doped Al 2 O 3 model cluster. The creep rate was correlated with the value of net charge in oxygen anion. The net charge of oxygen anion was one of the most important factors to determine the creep resistance in Al 2 O 3 .

Effects of Cooperativity in Proton Binding on the Net Charge of Proteins in Charge Ladders

This paper addresses the effects of cooperativity of proton binding on values of net charge and the change in net charge of a protein ΔZ due to the modification of charged groups. Capillary electrophoresis (CE) and charge ladderscollections of protein derivatives that differ in chargeare used to measure the net charge of lysozyme and values of pKa of the N-terminal α-amino group of this protein with different numbers of Lys e-amino groups acetylated. Values of pKa serve as local thermodynamic reporters of electrostatic potentials and therefore provide a direct measure of cooperativity in proton binding. Measured values are compared with values calculated using a model which combines the Poisson−Boltzmann (PB) equation and atomically detailed models of the distribution of charges on lysozyme with Monte Carlo (MC) sampling of different protonation states of the protein to determine the average extent of protonation of each titratable residue as a function of pH. Calculations with the PB-MC model together wi...

Vacancy effect of dopant cation on the high-temperature creep resistance in polycrystalline Al 2O 3

High-temperature creep resistance in polycrystalline Al 2O 3 with 0.1 mol% oxides of YO 1.5, ZrO 2 or MgO has been examined by uniaxial compression creep testing at 1250 °C. The creep resistance is highly improved by the doping of Y or Zr even in the dopant level of 0.1 mol%, but is retarded by Mg doping. The dopant effect on the creep resistance cannot be explained in terms of, for example, ionic radius of the dopant cation or eutectic point in Al 2O 3-oxide of dopant cation system. Each dopant cation was found to segregate in grain boundaries, and is likely to influence grain boundary diffusion in Al 2O 3. The ionic bonding and the covalent bonding of Al–O are lowered by the introduction of V O ′′ or V Al ′′′ but the values of the net charge in Al and O are increased by the cations doping. The change in the value of Net Charge is correlated well with the high-temperature creep resistance in Al 2O 3 with cation doping. It is suggested that the ionicity in Al and O is an important factor to determine high-temperature creep resistance in polycrystalline Al 2O 3.

Correlations Between the Charge of Proteins and the Number of Ionizable Groups They Incorporate: Studies Using Protein Charge Ladders, Capillary Electrophoresis, and Debye−Hückel Theory

The values of electrophoretic mobility, μelectro, of bovine carbonic anhydrase II, human carbonic anhydrase II, cytochrome c, lysozyme, superoxide dismutase, ovalbumin, and derivatives of these proteins produced by partial neutralization of Lys e-NH3+ and/or Asp and Glu carboxyl groups were measured using capillary electrophoresis (CE). For derivatives of these proteins with the lowest overall values of net charge (either positive or negative), the values of μelectro and the values of charge measured by CE, ZCE, demonstrate a linear correlation with the number of charged groups, n, converted to neutral derivatives. For derivatives of these proteins with larger values of net charge, the values of μelectro and ZCE demonstrate a nonlinear correlation with n. Several observations made in this work suggest that shifts in the values of pka of the ionizable groups on these proteins likely contribute to the observed nonlinear correlation. Debye−Huckel theory was used to calculate values of electrostatic potential...

Studies about Thermal Decarboxylation of Hydroxyl Benzoic Acids used in Thermal Imaging.

Six isomers of dihydroxyl benzoic acid and three isomers of trihydroxyl benzoic acid were analysed systematically with thermogravimetric analysis. They, the starting temperatures (TS), the peak temperatures (TP) and the end temperatures (Te) in the decarboxylation reactions, were worked out based on the integral curves (TG) and the differential curves (DTG) in thermogravimetric analysis of these compounds. The carbon atoms of benzene rings connected with carboxyl groups of these compounds are numbered as C1, and the carbon atoms of carboxyl groups are numbered as C7. The chemical shift values of C1 (δ1) and C7 (δ7) are from the NMR spectra of these compounds, and it is found that there is a linear relation between TP and δ1 or (δ7-δ1). The net charge values of C1 (q1) and C7 (q7) are from the calculations of quantum chemistry. After compared δ1 and(δ7-δ1) with q1 and(q7-q1) respectively, it is also found that there are linear relations between them. These prove that there is a close relation among δi, qi and the reactivity of i-position on the benzene ring of a phenolic compound.

Calculated potential for water enhanced crack growth in silicon

Ab initio molecular orbital calculations have been performed on the model molecules Si 8H 17OH (taking into consideration of the crack wall species -OH and -H) for the crack-tip strain simulation study of silicon and its reaction with water. The calculated equilibrium structure of silicon using a 6-31G ∗∗ basis set agreed well with experimental data. It was found that, regardless of the manner strain was applied, the values of net charge on the two crack-tip Si atoms are similar and that both become slightly more positive as strain increases. In addition to this absence of polarization of the crack-tip SiSi bond under strain, steric hindrance also inhibited the approach of the water molecule to the SiSi crack tip region. These calculations explain why moisture does not enhance crack-growth via chemical reaction in Si, which is different from that observed in silica.

A possible explanation for the photochromism of two 4-alkenyl-sydnones

Blue reversible photochromism of 3-phenyl-4-(3-methyl-1-buten-2-yl)sydnone IV and 3-phenyl-4-(1-cyclohexen-1-yl)sydnone V may be interpreted by an electrocyclic transformation in the sydnone ring leading to the N-nitrosoaminoketene system. Both sydnones are photochromic in frozen solutions. In the solid state only sydnone V is photochromic; stabilization of the coloured form is probably due to the intramolecular hydrogen bond between the carbonyl oxygen of the opened sydnone skeleton and hydrogen of the cyclohexene substituent. By a comparison between the experimentally recorded UV—VIS spectra and the respective theoretical spectra, obtained by the semiempirical MO INDO method, the validity of the ring opening model for sydnone photochromism is discussed. The net charge values in the ground and excited states suggest that the idea of the sydnone ring opening by cleavage of the O(1)C(5) bond could be justifiable.

Point-Discharge-Current Observations in the Thunderstorm Environment of the Years 1987 and 1988 at Pune

Observations of point-discharge current through an artificially erected single point in the thunderstorm environment at Pune during the years 1987 and 1988 were made and studied for the monthly net charge received by the earth through this process. Curiously, the net charge received by the earth at the station through point-discharge current, during six thunderstorm days in May and five in June 1987, was noticed as positive and amounted to 9 and 3 mC, respectively, whereas during the rest of 1987 and most of the time throughout 1988 the net-charge values were negative. A possible explanation of this observed feature is sought in an examination of the upper-atmospheric meteorological and electrical conditions that are reported responsible elsewhere and perhaps also in the present case.