Marta B. Peirotti

Hydration, charge, size, and shape characteristics of peptides from their CZE analyses.

A CZE model is presented for peptide characterization on the basis of well-established physicochemical equations. The effective mobility is used as basic data in the model to estimate relevant peptide properties such as, for instance, hydration, net and total electrical charge numbers, hydrodynamic size and shape, particle average orientation, and pH-microenvironment from the charge regulation phenomenon. Therefore 102 experimental effective mobilities of different peptides are studied and discussed in relation to previous work. An equation for the estimation of peptide hydration as a function of ionizing, polar, and non-polar amino acid residues is included in the model. It is also shown that the shape-orientation factor of peptides may be either lower or higher than one, and its value depends on a complex interplay among total charge number, molar mass, hydration, and amino acid sequence.

Estimation of global structural and transport properties of peptides through the modeling of their CZE mobility data

Peptide electrophoretic mobility data are interpreted through a physicochemical CZE model, providing estimates of the equivalent hydrodynamic radius, hydration, effective and total charge numbers, actual ionizing pK, pH-near molecule and electrical permittivity of peptide domain, among other basic properties. In this study, they are used to estimate some peptide global structural properties proposed, providing thus a distinction among different peptides. Therefore, the solvent drag on the peptide is obtained through a characteristic friction power coefficient of the number of amino acid residues, defined from the global chain conformation in solution. As modeling of the effective electrophoretic mobility of peptides is carried out in terms of particle hydrodynamic size and shape coupled to hydration and effective charge, a packing dimension related to chain conformation within the peptide domain may be defined. In addition, the effective and total charge number fractions of peptides provide some clues on the interpretation of chain conformations within the framework of scaling laws. Furthermore, the model estimates transport properties, such as sedimentation, friction and diffusion coefficients. As the relative numbers of ionizing, polar and non-polar amino acid residues vary in peptides, their global structural properties defined here change appreciably. Needs for further research are also discussed.

Analysis of the interplay among charge, hydration and shape of proteins through the modeling of their CZE mobility data

Electrophorectic mobility data of four proteins are analyzed and interpreted through a physicochemical CZE model, which provides estimates of quantities like equivalent hydrodynamic radius (size), effective charge number, shape orientation factor, hydration, actual pK values of ionizing groups, and pH near molecule, among others. Protein friction coefficients are simulated through the creeping flow theory of prolate spheroidal particles. The modeling of the effective electrophoretic mobility of proteins requires consideration of hydrodynamic size and shape coupled to hydration and effective charge. The model proposed predicts native protein hydrations within the range of values obtained experimentally from other techniques. Therefore, this model provides consistently other physicochemical properties such as average friction and diffusion coefficients and packing fractal dimension. As the pH varies from native conditions to those that are denaturing the protein, hydration and packing fractal dimension change substantially. Needs for further research are also discussed and proposed.

Global conformations of proteins as predicted from the modeling of their CZE mobility data.

Estimations of protein global conformations in well‐specified physicochemical microenvironments are obtained through global structural parameters defined from polypeptide‐scale analyses. For this purpose protein electrophoretic mobility data must be interpreted through a physicochemical CZE model to obtain estimates of protein equivalent hydrodynamic radius, effective and total charge numbers, hydration, actual ionizing pK and pH‐near molecule. The electrical permittivity of protein domain is also required. In this framework, the solvent drag on proteins is obtained via the characteristic friction power coefficient associated with the number of amino acid residues defining the global chain conformation in solution. Also, the packing dimension related to the spatial distribution of amino acid residues within the protein domain is evaluated and discussed. These scaling coefficients together with the effective and total charge number fractions of proteins provide relevant interpretations of protein global conformations mainly from collapsed globule to hybrid chain regimes. Also, protein transport properties may be estimated within this framework. In this regard, the central role played by the friction power coefficient in the evaluation of these properties is highlighted.

Evaluation of the slip length in the slipping friction between background electrolytes and peptides through the modeling of their capillary zone electrophoretic mobilities

This work analyzes and discusses several physicochemical peptide chain properties that may generate partial or total BGE slip boundary conditions on the surface of peptides migrating as spherical and aspherical particles in CZE. A definition of the BGE slip length is presented that is able to account the effect of particle curvature through the associated metrical coefficients. This definition allows the distinction between partial and total BGE slip lengths. It is also shown that the BGE slip length must be variable on orthotropic aspherical particles surfaces.

Interplay between electrophoretic mobility and intrinsic viscosity of polypeptide chains.

The present work is motivated specifically by the need to find a simple interplay between experimental values of electrophoretic mobility and intrinsic viscosity (IV) of polypeptides. The connection between these two properties, as they are evaluated experimentally in a formulated dilute solution, may provide relevant information concerning the physicochemical characterization and separation of electrically charged chains such as polypeptides. Based on this aspect, a study on the relation between the effective electrophoretic mobility and the IV of the following globular proteins is carried out: bovine carbonic anhydrase, staphylococcal nuclease, human carbonic anhydrase, lysozyme, human serum albumin. The basic interpretation of the IV through polypeptide chain conformations involves two unknowns: one is the Flory characteristic ratio involving short‐range intramolecular interactions and the other is the Mark–Houwink exponent associated with large‐range intramolecular interactions. Here, it will be shown via basic and well‐established electrokinetic theories and scaling concepts that the IV and global chain flexibility of polypeptides in dilute solutions may be estimated from capillary zone electrophoresis, in addition to classical transport properties. The polypeptide local chain flexibility may change due to electrostatic interactions among closer chain ionizing groups and the hindrance effect of their associated structural water.

Estimation of electrokinetic and hydrodynamic global properties of relevant amyloid‐beta peptides through the modeling of their effective electrophoretic mobilities and analysis of their propensities to aggregation

Neuronal activity loss may be due to toxicity caused by amyloid-beta peptides forming soluble oligomers. Here amyloid-beta peptides (1-42, 1-40, 1-39, 1-38, and 1-37) are characterized through the modeling of their experimental effective electrophoretic mobilities determined by a capillary zone electrophoresis method as reported in the literature. The resulting electrokinetic and hydrodynamic global properties are used to evaluate amyloid-beta peptide propensities to aggregation through pair particles interaction potentials and Brownian aggregation kinetic theories. Two background electrolytes are considered at 25°C, one for pH 9 and ionic strength I = 40 mM (aggregation is inhibited through NH4OH) the other for pH 10 and I = 100 mM (without NH4OH). Physical explanations of peptide oligomerization mechanisms are provided. The effect of hydration, electrostatic, and dispersion forces in the amyloidogenic process of amyloid-beta peptides (1-40 and 1-42) are quantitatively presented. The interplay among effective charge number, hydration, and conformation of chains is described. It is shown that amyloid-beta peptides (1-40 and 1-42) at pH 10, I = 100 mM and 25°C, may form soluble oligomers, mainly of order 2 and 4, after an incubation of 48 h, which at higher times evolve and end up in complex structures (protofibrils and fibrils) found in plaques associated with Alzheimers disease.

Determination of electrokinetic and hydrodynamic parameters of proteins by modeling their electrophoretic mobilities through the electrically charged spherical soft particle.

This work explores the possibility of using the electrically charged “spherical soft particle” (SSP) to model the electrophoretic mobility of proteins in the low charge regime. The general framework concerning the electrophoretic mobility of the SSP already presented in the literature is analyzed and discussed here in particular for polyampholyte‐polypeptide chains. In this regard, this theory is applied to BSA for different protocol pH values. The physicochemical conditions required to model proteins as SSP from their experimentally determined electrophoretic mobilities are established. In particular, the protein charge regulation phenomenon and the SSP particle core are included to study BSA having isoelectric point pI ≈ 5.71, within a wide range of bulk pH values. The results of this case study are compared with previous ones concerning the spherical porous particle and the spherical hard particle with occluded water. A discussion of chain conformations in the SSP polyampholyte layer is presented through estimations of the packing and friction fractal dimensions.

Global chain properties of an all l-α-eicosapeptide with a secondary α-helix and its all retro d-inverso-α-eicosapeptide estimated through the modeling of their CZE-determined electrophoretic mobilities.

Several global chain properties of relatively long peptides composed of 20 amino acid residues are estimated through the modeling of their experimental effective electrophoretic mobilities determined by CZE for 2 < pH < 6. In this regard, an all l‐α‐eicosapeptide, including a secondary α‐helix (Peptide 1) and its all retro d‐inverso‐α‐eicosapeptide (Peptide 2), are considered. Despite Peptides 1 and 2 are isomeric chains, they do not present similar global conformations in the whole range of pH studied. These peptides may also differ in the quality of BGE components chain interactions depending on the pH value. Three Peptide 1 fragments (Peptides 3, 4, and 5) are also analyzed in this framework with the following purposes: (i) visualization of the effects of initial and final strands at each side of the α‐helix on the global chain conformations of Peptide 1 at different pHs and (ii) analysis of global chain conformations of Peptides 1 and 2, and Peptide 1 fragments in relation to their pI values. Also, the peptide maximum and minimum hydrations predicted by the model, compatible with experimental effective electrophoretic mobilities at different pHs, are quantified and discussed, and needs for further research concerning chain hydration are proposed. It is shown that CZE is a useful analytical tool for peptidomimetic designs and purposes.

The Linear Viscoelastic Relaxation Modulus Related to the MWD of Linear Homopolymer Blends

This work analyzes the relation between the shear relaxation modulus of entangled, linear and flexible homopolymer blends and molecular weight distribution (MWD). The theory employed is based on the double reptation mixing rule and a law for the relaxation time of chains in a polydisperse matrix. It is shown that chain reptation with contour length fluctuations and tube constraint release are the relevant mechanisms of chain relaxation, when the polydispersity is high. The effect of the last mechanism is to decrease chain relaxation times. Calculations are carried out with rheometric data of linear viscoelasticity for polystyrene, polybutadiene, polypropylene and high density polyethylene. The model predictions are compared with experimental data of gel permeation chromatography (GPC). In homopolymer blends with a wide molecular weight distribution, only chains of relatively low molecular weight relax by reptation with contour length fluctuations. A relaxation law for chains in a polydisperse polymer matrix is suggested and validated with GPC data. The theory verifies that the zero-shear rate viscosity of a polydisperse polymer, is proportional to the mass average molecular weight raised to a power greater than that found for the near monodisperse counterpart.