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首页> 外文期刊>Molecular and Cellular Biology >Pentadecamer-binding proteins: definition of two independent protein-binding sites needed for functional activity.
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Pentadecamer-binding proteins: definition of two independent protein-binding sites needed for functional activity.

机译：Pentadecamer结合蛋白：功能活性所需的两个独立的蛋白结合位点的定义。

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The SP6 kappa-promoter pentadecamer (pd) element was found to be unable to stimulate transcription when present in one copy as the only promoter element in a minimal promoter but showed weak stimulatory activity when present as a multimer (four copies). One copy of the pd element acted synergistically with an octamer element, but not with a SP1 site, to stimulate transcription. The effect was orientation dependent with regard to the pd element. Gel shift analysis showed that pd-binding proteins were expressed in transformed as well as nontransformed B lymphocytes, irregardless of their differentiation stage, and in HeLa cells. Two major complexes, binding to different sites within the pd element, were observed in gel shifts. A low-molecular-weight form dominated in resting cells, while a higher-molecular-weight form appeared after mitogenic stimulation. Southwestern analysis showed that the low-molecular-weight pd-binding protein had a molecular mass of 35 kDa, which was confirmed by fractionation by denaturating polyacrylamide gel electrophoresis and molecular sieving. The higher-molecular-weight complex was sensitive to detergent treatment, while the low-molecular-weight complex was not. Mutation analysis showed that the two pd-binding complexes interacted with distinct sites within the element and that dual occupancy was required for functional activity. The functional synergy between the pd element and the octamer was more pronounced in plasmacytomas than in B-cell lymphomas.
机译：发现当一个拷贝中的SP6κ启动子pentadecamer（pd）元件作为最小启动子中的唯一启动子元件存在时，不能刺激转录，而当以多聚体形式（四个拷贝）存在时，则显示出弱的刺激活性。 pd元素的一个副本与八聚体元素（而不是SP1位点）协同作用，以刺激转录。关于pd元素，该效果取决于取向。凝胶位移分析表明，pd结合蛋白在转化的和未转化的B淋巴细胞中均表达，无论其分化阶段如何，都在HeLa细胞中表达。在凝胶位移中观察到两种主要的复合物，它们结合到pd元件内的不同位点。低分子形式在静息细胞中占主导，而有丝分裂刺激后出现高分子量形式。西南分析显示，低分子量pd结合蛋白的分子量为35 kDa，这是通过变性聚丙烯酰胺凝胶电泳和分子筛分馏来证实的。较高分子量的复合物对去污剂处理敏感，而低分子量的复合物对洗涤剂处理不敏感。突变分析表明，两个pd结合复合物与元素内的不同位点相互作用，功能活动需要双重占用。 pd元素和八聚体之间的功能协同作用在浆细胞瘤中比在B细胞淋巴瘤中更为明显。

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