HEME STAINING IN GELS, A USEFUL TOOL IN THE STUDY OF BACTERIAL C-TYPE CYTOCHROMES
HEME STAINING IN GELS, A USEFUL TOOL IN THE STUDY OF BACTERIAL C-TYPE CYTOCHROMES
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DOI：
10.1016/0005-2728(86)90234-3
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发表时间：
1986-12-03
期刊：
影响因子：
--
通讯作者：
PETTIGREW, GW
中科院分区：
文献类型：
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作者：
GOODHEW, CF;BROWN, KR;PETTIGREW, GW
C型细胞色素是唯一一种在十二烷基硫酸钠凝胶电泳法中能定量保留其血红素的细胞色素，可以通过它们的血红素过氧化物酶活性在复杂的混合物中进行鉴定。虽然较弱的染色条带可能是由于残留的血红素附着在b型细胞色素上或由于血红素的迁移，但这些影响可以通过预先用有机溶剂提取来消除。血红素染色的产色率可以估计特定细胞色素的相对数量，特别是在负载低于50pmoL的情况下。在较大负荷下，观察到颜色显影的平台期。新制成的凝胶颜色显影要差得多。血红素染色方法在细菌呼吸研究的三个特殊领域中被证明是有用的。首先，它可以评估革兰氏阴性细菌球体形成的结果。其次，血红素染色的定量有助于研究生长条件对C型细胞色素的诱导作用。第三，通过使用血红素染色的十二烷基硫酸钠电泳胶，大大简化了复杂的色谱图的解释。
c-Type cytochromes are the only cytochromes to retain quantitatively their haem during SDS gel electrophoresis and can be identified in complex mixtures by their haem peroxidase activity. Although weak staining bands may be due to residual haem attachment to b-type cytochromes or to migration of haem, these effects could be abolished by prior extraction with organic solvent. The colour yield of haem staining allowed an estimate of the relative amounts of a particular cytochrome, particularly if loadings were below 50 pmol. At greater loadings, a plateau of colour development was observed. Freshly made gels gave much poorer colour development. The haem-staining method was shown to be useful in three particular areas of study in bacterial respiration. Firstly, it allows assessment of the results of sphaeroplast formation in gram negative bacteria. Secondly, quantitation of the haem stain was useful in the investigation of the induction effects of growth conditions on c-type cytochromes. Thirdly, the interpretation of complex chromatographic profiles was greatly simplified by the use of haem-stained SDS electrophoretic gels.