Influence of Acetyl and Amide Groups on Peptides RP-LC Retention Behavior

̧czek, Tomasz Ba; Sieradzka, Magdalena

doi:10.1080/10826070802319438

Cited by 3 publications

(2 citation statements)

References 18 publications

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“…Recent studies by other groups investigated the influence of post-translational modifications [41,42]. Recent studies by other groups investigated the influence of post-translational modifications [41,42].…”

Section: Prediction Of Peptide Retention Timesmentioning

confidence: 99%

Peptide Purification by Reversed‐Phase Chromatography

Kusebauch¹,

McBee²,

Bletz³

et al. 2010

Amino Acids, Peptides and Proteins in Organic Chemistry

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Reversed-phase high-performance liquid chromatography (RP-HPLC) has dominated peptide purification since the late 1970s due to its efficient separation and ease of use with volatile compatible buffer systems for numerous applications [1,2]. RP-HPLC separation of chains of amino acids (peptides and proteins) is based predominantly on reversible hydrophobic interactions between the amino acid side chains with the hydrophobic surface of the chromatographic stationary phase (solid particles or packings) in competition with the flowing mobile phase (chromatographic buffer or solvent). Separation is performed by binding peptides onto the hydrophobic RP stationary phase in polar conditions (e.g., water) and eluting in nonpolar solvents (e.g., organic solvent such as acetonitrile) in the presence of a pH modifying agent. As peptides bind to the stationary phase, the amount of hydrophobic area on the surface of the peptide(s) exposed to the solvent is minimized. Thus, the degree of organized water is decreased with a concomitant favorable increase in entropy of the system. For this reason, it is advantageous, under these solvent conditions, for peptides to associate with the stationary phase upon loading onto the RP-HPLC column. Mobile phase composition is then subsequently modified so that bound peptides are differentially eluted (desorbed) back into the mobile phase. The order of peptide desorption is simply based on their relative hydrophobicity (i.e., least hydrophobic proteins elute first, followed by peptides in increasing order of heir surface hydrophobicity). Peptide elution is usually performed by increasing the organic solvent concentration (either in a stepwise fashion or in a gradient manner). By these means peptides, which are concentrated or trace enriched during the binding and separation process, are eluted in a purified and concentrated form ready for collection or subsequent analysis using hyphenated techniques such as mass spectrometry.

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Section: Prediction Of Peptide Retention Timesmentioning

confidence: 99%

Peptide Purification by Reversed‐Phase Chromatography

Kusebauch¹,

McBee²,

Bletz³

et al. 2010

Amino Acids, Peptides and Proteins in Organic Chemistry

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show abstract

“…Other studies associated with the influence of PTM on peptide retention was performed for the peptides possessing acetyl and/or amide groups [43]. Generally, the studies indicated that addition of amide group decreased and addition of acetyl group increased retention of peptides.…”

Section: Amino Acid Composition As a Core Basis For Predictive Modelsmentioning

confidence: 99%

Predictions of peptides' retention times in reversed‐phase liquid chromatography as a new supportive tool to improve protein identification in proteomics

Bączek

Kaliszan

2009

Proteomics

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One of the initial steps of proteomic analysis is peptide separation. However, little information from RP-HPLC, employed for peptides separation, is utilized in proteomics. Meanwhile, prediction of the retention time for a given peptide, combined with routine MS/MS data analysis, could help to improve the confidence of peptide identifications. Recently, a number of models has been proposed to characterize quantitatively the structure of a peptide and to predict its gradient RP-HPLC retention at given separation conditions. The chromatographic behavior of peptides has usually been related to their amino acid composition. However, different values of retention coefficients of the same amino acid in different peptides at different neighborhoods were commonly observed. Therefore, specific retention coefficients were derived by regression analysis or by artificial neural networks (ANNs) with the use of a set of peptides retention. In the review, various approaches for peptide elution time prediction in RP-HPLC are presented and critically discussed. The contribution of sequence dependent parameters (e.g., amphipathicity or peptide sequence) and peptide physicochemical descriptors (e.g., hydrophobicity or peptide length) that have been shown to affect the peptide retention time in LC are considered and analyzed. The predictive capability of the retention time prediction models based on quantitative structure-retention relationships (QSRRs) are discussed in details. Advantages and limitations of various retention prediction strategies are identified. It is concluded that proper processing of chromatographic data by statistical learning techniques can result in information of direct use for proteomics, which is otherwise wasted.

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Prediction of peptide retention time in reversed-phase liquid chromatography and its application in protein identification

Liu

Wang²,

Fu³

et al. 2010

Chinese Journal of Chromatography

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Liquid chromatography-mass spectrometry (LC-MS) is the mainstream of high throughput protein identification technology. Peptide retention time in reversed-phase liquid chromatography (RPLC) is mainly determined by the physicochemical properties of the peptide and the LC conditions (stationary phase and mobile phase). Retention time can be predicted by analyzing these properties and quantifying their effects on peptide chromatographic behavior. Prediction of peptide retention time in LC can be used to improve identification of peptides and post translational modifications (PTM). There are mainly two methods to predict retention time: i.e., retention coefficients and machine learning. The coefficient of determination between observed and predicted retention times can reach 0.93. With the development of LC-MS technology, retention time prediction will become an important tool to facilitate protein identification.

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Influence of Acetyl and Amide Groups on Peptides RP-LC Retention Behavior

Cited by 3 publications

References 18 publications

Peptide Purification by Reversed‐Phase Chromatography

Peptide Purification by Reversed‐Phase Chromatography

Predictions of peptides' retention times in reversed‐phase liquid chromatography as a new supportive tool to improve protein identification in proteomics

Prediction of peptide retention time in reversed-phase liquid chromatography and its application in protein identification

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