Application of hydrophilic-interaction liquid chromatography to the separation of phosphorylated H1 histones

Lindner, Herbert; Sarg, Bettina; Helliger, Wilfried

doi:10.1016/s0021-9673(97)00468-8

Cited by 92 publications

(66 citation statements)

References 41 publications

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“…Hydrophilic-interaction LC (HILIC) is a powerful technique that has proved to be extremely well suited for histone separation. Lindner optimized HILIC to attain excellent resolution of core and linker histones [97][98][99][100]. In their work, the protein mixture was fractionated on a weak cation-exchange column using an increasing sodium perchlorate gradient in a methanephosphonic acid-triethylamine buffer (pH 3.0) in the presence of organic solvent.…”

Section: Hydrophilic-interaction Lc and Offline Hydrophilic Interactionmentioning

confidence: 99%

Mass spectrometry-based strategies for characterization of histones and their post-translational modifications

Ren

Freitas

2007

Expert Review of Proteomics

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Due to the intimate interactions between histones and DNA, the characterization of histones has become the focus of great attention. A series of mass spectrometry-based technologies have been dedicated to the characterization and quantitation of different histone forms. This review focuses on the discussion of mass spectrometry-based strategies used for the characterization of histones and their post-translational modifications. Keywordshistone; mass spectrometry; post-translational modification Histones are a group of highly conserved proteins throughout eukaryotic evolution [1]. The core histones (H4, H3, H2A and H2B) form an octamer, in which a H2A-H2B dimer associates on each side of the H3-H4 tetramer through an interaction between the C-terminal halves of H2B and H4 [2]. The negatively charged DNA superhelix wraps around the histone octamer to form repeating nucleosome cores, which further assemble into higher order chromatin fibers stabilized by the linker histones (H1 and H5) and linker DNA [3]. The histone fold regions in core histones are highly conserved α-helices, which are connected by two loops. The third histone structural motif comprises the flexible and irregular histone tails, which are extended and reach outside of the nucleosomal unit to interact with DNA [4]. Challenge of molecular characterization: post-translational modifications & sequence variationsAs a building block of the nucleosome, histones play an important role in chromatin assembly and affect the interactions between DNA and other chromatin-associated proteins. Histones regulate gene transcription through their diverse post-translational modifications (PTMs), which include lysine acetylation, lysine and arginine methylation, serine and threonine phosphorylation, lysine ubiquitination, lysine sumoylation, and poly-ADP-ribosylation of glutamic acid [5][6][7][8]. Among all the PTMs, histone acetylation and methylation have been most †

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Section: Hydrophilic-interaction Lc and Offline Hydrophilic Interactionmentioning

confidence: 99%

Mass spectrometry-based strategies for characterization of histones and their post-translational modifications

Ren

Freitas

2007

Expert Review of Proteomics

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“…Bonenfant et al applied C18 RP LC-MS to the study of histones derived from Jurkat cells [16]. Hydrophilic-interaction liquid chromatography (HILIC) coupling with off-line mass spectrometry has been successful in separating and characterizing different histone isoforms [18,19]. However, the high salt required to elute the histones adds an additional desalting step prior to MS analysis.…”

Section: Introductionmentioning

confidence: 99%

Liquid chromatography mass spectrometry profiling of histones

Jacob

Amunugama

et al. 2007

Journal of Chromatography B

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Here we describe the use of reverse-phase liquid chromatography mass spectrometry (RP-LC-MS) to simultaneously characterize variants and post-translationally modified isoforms for each histone. The analysis of intact proteins significantly reduces the time of sample preparation and simplifies data interpretation. LC-MS analysis and peptide mass mapping have previously been applied to identify histone proteins and to characterize their post-translational modifications. However, these studies provided limited characterization of both linker histones and core histones. The current LC-MS analysis allows for the simultaneous observation of all histone PTMs and variants (both replacement and bulk histones) without further enrichment, which will be valuable in comparative studies. Protein identities were verified by the analysis of histone H2A species using RPLC fractionation, AU-PAGE separation and nano-LC-MS/MS.

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“…Los resultados obtenidos se contrastaron con distintos protocolos de separación realizados por varios investigadores (42,(48)(49)(50). En uno de los trabajos, el proceso de separación de las histonas H1 de hígado de rata se realizó durante 45 min a un flujo de 0,5 ml/min, con gradiente lineal desde 60% de disolvente A y 40% de disolvente B hasta 42% de A y 58% de B (disolvente A: 0,1% de ácido trifluoroacético en agua; disolvente B: 0,1% de ácido trifluoroacético en 70% de acetonitrilo) y obtuvieron 6 picos, correspondientes a las histonas H1°, H1b, H1a, H1d, H1e+c y H1c (48).…”

Section: Purificación De Los Subtiposunclassified

“…Con esta cromatografía, las histonas se separaron en dos picos: en el primero, se separó la H1b, y el segundo correspondió a una mezcla de los otros subtipos. Para separar los subtipos de la mezcla, se utilizó una cromatografía líquida de interacción hidrofílica (49).…”

Section: Purificación De Los Subtiposunclassified

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Caracterización de la estructura secundaria de subtipos de la histona H1 por dicroísmo circular

2015

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Application of hydrophilic-interaction liquid chromatography to the separation of phosphorylated H1 histones

Cited by 92 publications

References 41 publications

Mass spectrometry-based strategies for characterization of histones and their post-translational modifications

Mass spectrometry-based strategies for characterization of histones and their post-translational modifications

Liquid chromatography mass spectrometry profiling of histones

Caracterización de la estructura secundaria de subtipos de la histona H1 por dicroísmo circular

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