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Author:
Date:
2015-08-05
[Abstract] Among the seven serines and one threonine in the carboxyl-terminus of HBV C protein, all but one (serine 183) appear in the context of RxxS/T consensus phosphoacceptor motifs and also overlap with other consensus motifs, such as S/TP, RS, SPRRR, RRRS/T, or RRxS/T, suggesting that various cellular kinases phosphorylate these residues. To determine whether threonine and/or serine (serines 157, 164, 170, 172, 178, and 180, and threonine 162, adw subtype) of HBV C protein are indeed phosphoacceptor residues in cells, Huh7 were transfected with a series of C-protein-expressing mutants, labeled ...
[摘要] 在HBV C蛋白的羧基末端中的七个丝氨酸和一个苏氨酸中，除了一个（丝氨酸183）外，所有其它丝氨酸出现在RxxS/T共有磷酸受体基序的上下文中，并且还与其他共有基序重叠，例如S/TP，RS ，SPRRR，RRRS/T或RRxS/T，表明各种细胞激酶磷酸化这些残基。 为了确定HBV C蛋白的苏氨酸和/或丝氨酸（丝氨酸157,164,170,172,178和180以及苏氨酸162，adw亚型）是否确实是细胞中的磷酸受体残基，Huh7用一系列C- 蛋白表达突变体，用正磷酸32P标记14小时，然后裂解。 用抗HBc抗体免疫沉淀 32 P标记的裂解物，并通过放射自显影检测32 P标记的免疫沉淀的C蛋白。
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Author:
Date:
2014-07-05
[Abstract] Pulse-chase method is a powerful technique used to follow the dynamics of proteins over a period of time. The expression level, processing, transport, secretion or half-life of proteins can be tracked by metabolically labeling the cells, such as with radiolabeled amino acids (pulse step). This protocol describes the condition used to study the folding and disulfide bond formation of immunoglobulin in suspension cells. With some minor modifications, this protocol can be adapted to study the degradation rate or the secretion of target proteins.
[摘要] 脉冲追踪法是用于跟踪蛋白质在一段时间内的动力学的有力技术。 蛋白质的表达水平，加工，转运，分泌或半衰期可以通过代谢标记细胞，例如用放射性标记的氨基酸（脉冲步骤）来追踪。 该方案描述了用于研究悬浮细胞中免疫球蛋白的折叠和二硫键形成的条件。 通过一些微小的修改，这个协议可以适应于研究目标蛋白的降解速率或分泌。
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