Hsp70 regulates the interaction between the peroxisome targeting signal type 1 (PTS1)-receptor Pex5p and PTS1
Hsp70 regulates the interaction between the peroxisome targeting signal type 1 (PTS1)-receptor Pex5p and PTS1
复制标题
DOI：
10.1042/0264-6021:3570157
复制
发表时间：
2001-07-01
影响因子：
4.1
通讯作者：
Fujiki, Y
中科院分区：
文献类型：
--
作者：
Harano, T;Nose, S;Fujiki, Y
四三肽重复序列 (TPR) 基序家族的过氧化物酶体靶向信号 1 型 (PTS1) 受体 Pex5p 主要位于细胞质中，介导 PTS1 蛋白易位至过氧化物酶体。作为了解蛋白质输入过氧化物酶体机制的一步，我们使用无细胞合成酰基辅酶A氧化酶（AOx）作为 PTSI 货物蛋白，研究了 Pex5p 识别 PTSI 所需的能量和胞质因子的分子机制。以及来自大鼠肝脏的 Pex5p 和热休克蛋白 (Hsp)70。 Pex5p与大鼠肝脏的过氧化物酶体部分相关，耐高浓度盐洗涤和碱提取，并且不能与外部添加的蛋白酶接触。 Pex5p 以 ATP 依赖性方式与 AOx 结合。将兔网织红细胞裂解物在无细胞翻译系统中合成的 AOx 导入过氧化物酶体中，无需补充 Pex5p 和 Hsp70，这意味着过氧化物酶体相关的 Pex5p 从膜中释放出来，并在此体外导入测定中发挥作用。针对 Pex5p 和 Hsp70 的抗体抑制 AOx 的输入。相比之下，在小麦胚芽裂解液中合成的 AOx 需要外部添加 Pex5p 才能导入。其中 Hsp70 增强了 AOx 的导入。 Pex5p 的 TPR 结构域以不依赖于 Hsp70 的方式与 N 末端部分结合，而在 Hsp70 存在的情况下注意到 TPR 区域的相互作用。 Hsp70 与 Pex5p 的 TPR 结构域相互作用。此外，Hsp70 和 ATP 协同增强 Pex5p 与 AOx 的 C 端 PTS 1 含有部分的结合，这意味着 Pex5p 在体内通过伴侣辅助和能量依赖机制识别其货物 PTS1 蛋白。
The peroxisome targeting signal type 1 (PTS1) receptor, Pex5p, of the tetratricopeptide repeat (TPR) motif family is located mostly in the cytosol and mediates the translocation of PTS1 proteins to peroxisomes. As a step towards understanding the mechanisms of protein import into peroxisomes, we investigated the molecular mechanisms involved in PTSI recognition by Pex5p with regard to requirement of energy and cytosolic factors, using cell-free synthesized acyl-CoA oxidase (AOx) as a PTSI cargo protein. together with Pex5p and heat-shock protein (Hsp)70 from rat liver. Pex5p was partly associated with peroxisomes of rat liver, was resistant to washing with a high concentration of salt and to alkaline extraction and was inaccessible to protease added externally. Pex5p bound to AOx in an ATP-dependent manner. AOx synthesized in a cell-free translating system from rabbit reticulocyte lysate was imported into peroxisomes without bring supplemented with Pex5p and Hsp70, implying that peroxisome-associated Pex5p was released from the membranes and functional in this in vitro import assay. Antibodies against Pex5p and Hsp70 inhibited AOx import. In contrast, AOx synthesized in a wheat-germ lysate required the external addition of Pex5p for import. in which Hsp70 augmented the AOx import. The TPR domain of Pex5p was revealed to bind to the N-terminal part in an Hsp70-independent manner, whereas mutual interaction of the TPR region was noted in the presence of Hsp70. Hsp70 interacted with the TPR domain of Pex5p. Moreover, Hsp70 and ATP synergistically enhanced the binding of Pex5p to the C-terminal PTS 1-containing part of AOx, implying that Pex5p recognizes its cargo PTS1 protein by chaperone assisted as well as energy-dependent mechanisms in vivo.