Behavior of peptides in capillary electrophoresis: Effect of peptide charge, mass and structure

Cifuentes, Alejandro; Poppe, H.

doi:10.1002/elps.1150181227

Cited by 98 publications

(97 citation statements)

References 139 publications

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“…The ''net charge concept'' is useful for predicting CZE migration behavior from the net charge and size ($mass) of an analyte [54]. In CEC, the situation is complicated by the fact that the mobile phase contains a significant amount of organic modifier.…”

Section: Net Charge and Electrophoretic Mobility Of The Peptidesmentioning

confidence: 99%

Electrochromatographic retention of peptides on strong cation‐exchange stationary phases

Nischang¹,

Höltzel²,

Tallarek³

2010

Electrophoresis

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We analyze the systematic and substantial electrical field-dependence of electrochromatographic retention for four counterionic peptides ([Met5]enkephalin, oxytocin, [Arg8]vasopressin, and luteinizing hormone releasing hormone (LHRH) ) on a strong cation-exchange (SCX) stationary phase. Our experiments show that retention behavior in the studied system depends on the charge-selectivity of the stationary phase particles, the applied voltage, and the peptides' net charge. Retention factors of twice positively charged peptides ([Arg8]vasopressin and LHRH at pH 2.7) decrease with increasing applied voltage, whereas lower charged peptides (oxytocin and [Met5]enkephalin at pH 2.7, [Arg8]vasopressin and LHRH at pH 7.0) show a concomitant increase in their retention factors. The observed behavior is explained on the basis of electrical fieldinduced concentration polarization (CP) that develops around the SCX particles of the packing. The intraparticle concentration of charged species (buffer ions, peptides) increases with increasing applied voltage due to diffusive backflux from the enriched CP zone associated with each SCX particle. For twice charged and on the SCX phase strongly retained peptides the local increase in mobile phase ionic strength reduces the electrostatic interactions with the stationary phase, which explains the decrease of retention factors with increasing applied voltage and CP intensity. Lower charged and weaker retained peptides experience a much stronger relative intraparticle enrichment than the twice-charged peptides, which results in a net increase of retention factors with increasing applied voltage. The CP-related contribution to electrochromatographic retention of peptides on the SCX stationary phase is modulated by the applied voltage, the mobile phase ionic strength, and the peptides' net charge and could be used for selectivity tuning in difficult separations.

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Section: Net Charge and Electrophoretic Mobility Of The Peptidesmentioning

confidence: 99%

Electrochromatographic retention of peptides on strong cation‐exchange stationary phases

Nischang¹,

Höltzel²,

Tallarek³

2010

Electrophoresis

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“…The chromatographic retention or electrophoretic migration time contains information on the physiochemical properties of the peptides, such as hydrophobicity in RPC and size and charge in standard capillary zone electrophoresis (CZE). A model of chromatographic retention [124][125][126][127][128] or electrophoretic migration [129][130][131][132][133][134][135][136][137][138] can be fitted to experimental data from known proteins. These models are then used to predict the retention time for candidate peptides in a database.…”

Section: Mass Spectrometry Peptide Mass Fingerprinting and Informationmentioning

confidence: 99%

“…Most models of electrophoretic mobility of peptides are non-linear functions of peptide charge and molecular weight [136]. The molecular weight is easily calculated for each peptide candidate, and the charge, or distribution of charges, is estimated from measured pKa values of amino acids in N and C-termini and side chain pKa values.…”

Section: Mass Spectrometry Peptide Mass Fingerprinting and Informationmentioning

confidence: 99%

“…In addition to the accurate mass measurement and distribution of peptides in the protein sequence, containing information on the non-random behaviour of trypsin, protein structure, post-translational modifications and database sequence errors, there is also information on the physiochemical properties of the individual peptides. This is primarily hydrophobicity in reversed-phase chromatography [124][125][126][127][128] and size and charge in capillary zone electrophoresis [129][130][131][132][133][134][135][136][137][138]. A predictor of retention time according to Eq.…”

Section: Figure 23 (Color Panel Opposite Side) Lc-fticr (A) and Ce-mentioning

confidence: 99%

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Identification and Characterization of Peptides and Proteins Using Fourier Transform Ion Cyclotron Resonance Mass Spectrometry

Palmblad

Bergquist

2003

Journal of Chromatography Library

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Mass spectrometry has in recent years been established as the standard method for protein identification and characterization in proteomics with excellent intrinsic sensitivity and specificity. Fourier transform ion cyclotron resonance is the mass spectrometric technique that provides the highest resolving power and mass accuracy, increasing the amount of information that can be obtained from complex samples. This thesis concerns how useful information on proteins of interest can be extracted from mass spectrometric data on different levels of protein structure and how to obtain this data experimentally. It was shown that it is possible to analyze complex mixtures of protein tryptic digests by direct infusion electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry and identify abundant proteins by peptide mass fingerprinting. Coupling on-line methods such as liquid chromatography and capillary electrophoresis increased the number of proteins that could be identified in human body fluids. Protein identification was also improved by novel statistical methods utilizing prediction of chromatographic behavior and the non-randomness of enzymatic digestion. To identify proteins by short sequence tags, electron capture dissociation was implemented, improved and finally coupled on-line to liquid chromatography for the first time. The combined techniques can be used to sequence large proteins de novo or to localize and characterize any labile post-translational modification. New computer algorithms for the automated analysis of isotope exchange mass spectra were developed to facilitate the study of protein structural dynamics. The non-covalent interaction between HIV-inhibitory peptides and the oligomerization of amyloid β-peptides were investigated, reporting several new findings with possible relevance for development of anti-HIV drug therapies and understanding of fundamental mechanisms in Alzheimer's disease.

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“…A number of reviews can be recommended in this field: * For peptide/protein separations, a series of journal reviews covering the field up to 1997 [7][8][9][10][11]. A more recent one [12], covers the period 1997-1999 for peptides, whereas for peptides/proteins of biotechnological interest, in the same time period, the field is surveyed by Lagu [13].…”

Section: Introductionmentioning

confidence: 99%

Capillary electrophoretic analysis of proteins and peptides of biomedical and pharmacological interest

Righetti

2001

Biopharm & Drug Disp

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Capillary electrophoresis (CE) is an automated approach to electrokinetic separations that has had a deep impact in all fields of life sciences, including biomedical and biotechnological research and clinical and forensic practice. The present review highlights aspects of peptides and proteins separations, with particular emphasis on macromolecular analytes of biomedical interest. Among the various CE techniques available, a novel methodology is here illustrated consisting in separations in acidic, isoelectric buffers, which have the advantage of protonating the silica wall, thus minimizing interactions of proteinaceous material with the siliceous surface, while allowing delivery of high voltage gradients, due to their low conductivities. The review ends with applications of CE to the analysis of folding/unfolding/refolding/misfolding of proteins, a field which has deep implications in the biomedical arena, since it is connected to a host of disorders, such as prion protein diseases.

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Behavior of peptides in capillary electrophoresis: Effect of peptide charge, mass and structure

Cited by 98 publications

References 139 publications

Electrochromatographic retention of peptides on strong cation‐exchange stationary phases

Electrochromatographic retention of peptides on strong cation‐exchange stationary phases

Identification and Characterization of Peptides and Proteins Using Fourier Transform Ion Cyclotron Resonance Mass Spectrometry

Capillary electrophoretic analysis of proteins and peptides of biomedical and pharmacological interest

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