ARP2 & 3 COMPLEX:REQD FOR ACTIN DEPEND MITOCHOND MOTILITY & INHERIT IN BUD YEAST

酵母ADA蛋白与TBP类SPT蛋白相互作用 多个复合体(ADA/SPT复合体) 转录的激活和抑制。我们已经确认了Tra1 作为通过串联质量的这些调节复合体的组成部分 Ngg1/Ada3相关基因的光谱分析。Tra1 是一组可能的蛋白激酶的成员，这些蛋白激酶有 与磷脂酰肌醇3-激酶相关的无羧基末端区域。 Tra1和ADA/SPT组件之间的相互作用通过 HA-Ada2和myc-Tra1的相互共沉淀 也含有Spt7的一个或多个复合体中的全细胞提取物。这个 提出了Tra1与多个ADA/SPT复合体的关联 通过与Ada2、Ngg1和Spt7在两个单独的峰中进行共分馏 在FPLC Mono Q柱上。在没有Ada2的情况下，洗脱曲线 TRA1P的峰位移至单一明显峰。Tra1还与 AdA/SPT蛋白在DNA-纤维素柱上结合Tra1 到DNA-纤维素需要ADA组分。以及允许 ADA/SPT络合物中Tra1p的初步测定 质谱学分析鉴定了一种位于 蛋白质的氨基末端。

The yeast ADA proteins and TBP-class of SPT proteins interact in multiple complexes (ADA/SPT complexes) that are required for the activation and repression of transcription. We have identified Tra1 as a component of these regulatory complexes through the tandem mass spectrometry analysis of proeints that associate with Ngg1/Ada3. Tra1 is a member of a group of putative protein kinases, that have acarboxyl-terminal region related to phosphatidylinisitol 3-kinases. The interaction between Tra1 and ADA/SPT components was verified by the reciprocal coimmiunprecipitation of HA-Ada2 and myc-Tra1 from whole cell extracts in one or complexes also containing Spt7. The association of Tra1 with more that one ADA/SPT complex was suggested by its cofractionation with Ada2, Ngg1 and Spt7 in two separate peaks on a FPLC mono Q column. In the absence of Ada2, the elution profile of Tra1p shifted to a single distinct peak. Tra1 also copurified with ADA/SPT proteins on a DNA-cellulose column where the binding of Tra1 to DNA-cellulose required the ADA components. As well as allowing the initial determination of Tra1p within the ADA/SPT complexes the tandem mass spectrometry analysis identified a phosphopeptide located near the protein's amino-terminus.