Rapid analysis of mitotic histone H1 phosphorylation by cationic disc electrophoresis at neutral pH in minigels

Paulson, James R.; Mesner, Peter W.; Delrow, Jeffrey J.; Mahmoud, Najwa; Ciesielski, Wayne A.

doi:10.1016/0003-2697(92)90307-s

Cited by 8 publications

(3 citation statements)

References 34 publications

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“…Note the presence of cytochrome P450 isoforms in the 2D gel. The first dimension, 16BAC gels were run either at pH 2 in the phosphate/glycine system [65] (panel A), or at pH 5 in the acetate/beta alanine system [80] (panel B), or at pH 7 in the Hepes/histidine system [81] (panel C). protein detection by silver ammonia staining [18] Note the increased streaking with increasing running pH…”

Section: Discussionmentioning

confidence: 99%

Fully denaturing two‐dimensional electrophoresis of membrane proteins: A critical update

et al. 2008

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The quality and ease of proteomics analysis depends on the performance of the analytical tools used, and thus of the performances of the protein separation tools used to deconvolute complex protein samples. Among protein samples, membrane proteins are one of the most difficult sample classes, because of their hydrophobicity and embedment in the lipid bilayers. This review deals with the recent progresses and advances made in the separation of membrane proteins by 2-DE separating only denatured proteins. Traditional 2-D methods, i.e., methods using IEF in the first dimension are compared to methods using only zone electrophoresis in both dimensions, i.e., electrophoresis in the presence of cationic or anionic detergents. The overall performances and fields of application of both types of method is critically examined, as are future prospects for this field.

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Section: Discussionmentioning

confidence: 99%

Fully denaturing two‐dimensional electrophoresis of membrane proteins: A critical update

et al. 2008

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“…A practical limitation to the actual use of these buffers for electrophoresis under native conditions is the sensitivity of several proteins to the exposure to acidic pH; loosening of H‐bonding pattern and structural rearrangements result in various phenomena, from loss of ligands/cofactors to decrease in solubility and aggregation. Discontinuous buffer systems for the electrophoresis of cationic proteins at near‐neutral pH were presented (K + as the leading ion, histidine as the trailing ion and either 3‐[ N ‐morpholine]propanesulfonic acid 75 or HEPES 76 as buffering counterion). One of them 76 has been applied in few instances 77–79 for the analysis of histones − more specifically, for the assessment of the extent of phosphorylation of H1 under various experimental conditions.…”

Section: Analysis Of High Pi Proteinsmentioning

confidence: 99%

“…Discontinuous buffer systems for the electrophoresis of cationic proteins at near‐neutral pH were presented (K + as the leading ion, histidine as the trailing ion and either 3‐[ N ‐morpholine]propanesulfonic acid 75 or HEPES 76 as buffering counterion). One of them 76 has been applied in few instances 77–79 for the analysis of histones − more specifically, for the assessment of the extent of phosphorylation of H1 under various experimental conditions. On the contrary, a cationic system combined to the presence of urea to provide resolution under dissociating/denaturing conditions has found extensive application in the format of acid‐urea (AU) and acid‐urea‐Triton (AUT) gels.…”

Section: Analysis Of High Pi Proteinsmentioning

confidence: 99%

Other than IPG‐DALT: 2‐DE variants

2010

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Special problems require special solutions. That is why, despite the trend towards standardization of the procedures in proteomic investigations, a number of alternative protocols, based on 2-DE, have been devised with time and are applied to the resolution of proteins on which the performance of IPG-DALT is poor. The difficult-to-handle samples include high pI, high molecular size and high hydrophobicity proteins. The protocol variants entail changes in the gel media and/or the use of unusual additives (often surfactants or solvents). But also: Specific questions require specific answers. When the aim is not the resolution of individual polypeptide chains in a fully unfolded state but the analysis of higher-order structures (proteins assembled from subunits, complexes assembled from interacting proteins) the standard conditions under which IPG-DALT is performed turn inadequate. Electrophoresis is then performed either in first dimension or in both first dimension and second dimension under non-denaturing (native) and/or non-reducing conditions.

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One‐ and two‐dimensional histone separations in acidic gels: Usefulness of methylene blue‐driven photopolymerization

1996

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We tested the recently introduced methylene blue-toluene sulfinate-diphenyl-iodonium polymerization system in order to prepare acetic acid-urea-Triton X-100 gels for histone separations. When compared to standard persulfate-based initiators, this system exhibited several advantages. First, the polymerization proceeds at a much faster rate but is easily controlled since it is light-dependent. Second, no prerunning of the gel was required, since there is no oxidizing molecule able to introduce artifacts. Moreover, this procedure produces gels presenting cleaner backgrounds with silver staining. The ability of the procedure to carry out high resolution two-dimensional electrophoresis of histones was also examined. High resolution two-dimensional gels, using photopolymerized acidic gels as the first or second dimension were obtained.

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Rapid analysis of mitotic histone H1 phosphorylation by cationic disc electrophoresis at neutral pH in minigels

Cited by 8 publications

References 34 publications

Fully denaturing two‐dimensional electrophoresis of membrane proteins: A critical update

Fully denaturing two‐dimensional electrophoresis of membrane proteins: A critical update

Other than IPG‐DALT: 2‐DE variants

One‐ and two‐dimensional histone separations in acidic gels: Usefulness of methylene blue‐driven photopolymerization

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