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首页> 外文期刊>Analytica chimica acta >Cu~(2+)-assisted two dimensional charge-mass double focusing gel electrophoresis and mass spectrometric analysis of histone variants
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Cu~(2+)-assisted two dimensional charge-mass double focusing gel electrophoresis and mass spectrometric analysis of histone variants

机译：Cu〜（2+）辅助二维电荷质量双聚焦凝胶电泳及组蛋白变异体的质谱分析

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Abundant isoforms and dynamic posttranslational modifications cause the separation and identification of histone variants to be experimentally challenging. To meet this need, we employ two-dimensional electrophoretic gel separation followed by mass spectrometric detection which takes advantage of the chelation of Cu~(2+) with amino acid residues exposed on the surfaces of the histone proteins. Acid-extracted rat liver histones were first mixed with CuSO4 solution and then separated in one dimension with triton-acid-urea (TAU) gel electrophoresis and in a second dimension using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The separations result from both the changes in charge and mass upon Cu~(2+) chelation. Identities of each separated gel bands were obtained by using matrix-assisted laser desorption-ionization mass spectrometry (MALDl-MS). It was found that the migration of H3 histone isoforms of rat liver is markedly affected by the use of Cu~(2+) ions.
机译：丰富的同工型和动态的翻译后修饰导致组蛋白变体的分离和鉴定在实验上具有挑战性。为了满足这一需求，我们采用二维电泳凝胶分离，然后进行质谱检测，该检测利用了Cu〜（2+）与组蛋白表面暴露的氨基酸残基的螯合作用。首先将酸提取的大鼠肝脏组蛋白与CuSO4溶液混合，然后使用triton-酸-尿素（TAU）凝胶电泳进行一维分离，然后使用十二烷基硫酸钠聚丙烯酰胺凝胶电泳（SDS-PAGE）进行第二维分离。分离是由于Cu〜（2+）螯合后电荷和质量的变化所致。通过使用基质辅助激光解吸电离质谱（MALD1-MS）获得每个分离的凝胶带的身份。发现Cu〜（2+）离子对大鼠肝脏H3组蛋白亚型的迁移有明显的影响。

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