SPECTROSCOPIC STUDIES OF CIBACRON BLUE AND CONGO RED BOUND TO DEHYDROGENASES AND KINASES - EVALUATION OF DYES AS PROBES OF THE DINUCLEOTIDE FOLD
SPECTROSCOPIC STUDIES OF CIBACRON BLUE AND CONGO RED BOUND TO DEHYDROGENASES AND KINASES - EVALUATION OF DYES AS PROBES OF THE DINUCLEOTIDE FOLD
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DOI：
10.1021/bi00590a026
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发表时间：
1979-01-01
期刊：
影响因子：
2.9
通讯作者：
WOODY, RW
中科院分区：
文献类型：
--
作者：
EDWARDS, RA;WOODY, RW
汽巴蓝和刚果红与核苷酸结合酶紧密结合，并且前一种染料已被证明可用于此类酶的亲和层析。当与酶结合时，这种染料的构象是令人感兴趣的，因为已经表明它们对酶的亲和力来自于模拟结合的辅因子的构象的能力。通过诱导圆二色性（CD）、结合测量以及与适当辅酶或底物的竞争，研究了汽巴蓝和刚果红与几种脱氢酶（牛乳酸脱氢酶、马和酵母醇脱氢酶）、激酶（猪腺苷酸激酶和酵母磷酸甘油酸激酶）和醛缩酶的相互作用。苯并红嘌呤4 B，一个二甲基类似物的刚果红和Cibacron蓝连接到葡聚糖或琼脂糖基质，用于检查的灵敏度的相互作用的扰动的染料结构。所考虑的染料不是核苷酸或辅酶的高度特异性类似物。这些染料在具有二核苷酸折叠的不同酶上不呈现单一独特的构象。诱导CD表明，对于密切相关的蛋白质或染料结构的小变化，结合染料的构象非常相似。尽管这些染料Cibacron Blue和刚果红缺乏独特的构象，但除了它们在亲和色谱中的有用性之外，它们还是核苷酸结合酶的有价值的构象探针。
Cibacron Blue and Congo Red bound tightly to nucleotide-binding enzymes, and the former dye has proven useful in affinity chromatography of such enzymes. The conformation of such dyes when bound to enzymes is of interest because it has been suggested that their affinity for an enzyme results from an ability to mimic the conformation of the bound cofactor. The interaction of Cibacron Blue and Congo Red with several dehydrogenases (bovine lactic dehydrogenase, equine, and yeast alcohol dehydrogenase) and kinases (porcine adenylate kinase and yeast phosphoglycerate kinase) and aldolase was studied by induced circular dichroism (CD), binding measurements and competition with the appropriate coenzymes or substrates. Benzopurpurin 4B, a dimethyl analogue of Congo Red and Cibacron Blue linked to dextran or Sepharose matrices were used to examine the sensitivity of the interaction to perturbations of the dye structure. The dyes under consideration are not highly specific analogues of nucleotides or coenzymes. These dyes do not assume a single unique conformation on different enzymes possessing the dinucleotide fold. The induced CD indicates that for closely related proteins or small variations in dye structure the conformation of the bound dye quite similar. Despite the lack of a unique conformation for these dyes Cibacron Blue and Congo Red are valuable conformational probes for nucleotide-binding enzymes, in addition to their usefulness in affinity chromatography.