Irmer H, Höhfeld J
Zentrum für Molekulare Biologie, Universität Heidelberg, Im Neuenheimer Feld 282, D-69120 Heidelberg, Germany.
J Biol Chem. 1997 Jan 24;272(4):2230-5. doi: 10.1074/jbc.272.4.2230.
The homo-oligomeric Hip protein cooperates with the 70-kDa heat shock cognate Hsc70 in the folding of newly synthesized polypeptide chains and in the conformational regulation of signaling molecules known to interact with Hsc70 and Hsp90. In order to further assess the role of Hip during protein biogenesis, a structure-function analysis of the Hip protein was initiated. By employing the yeast two-hybrid system, the Hsc70-binding site of Hip was mapped to a domain comprising multiple tetratricopeptide repeats and flanking charged alpha-helices. Affinity chromatography confirmed direct interaction of isolated Hip fragments and protein fusions bearing this region with the ATPase domain of Hsc70 in an ATP- and salt-dependent manner. Contact of Hip with the ATPase domain appears to be mediated primarily by the positively charged alpha-helix following the tetratricopeptide repeats. Furthermore, a domain required for homo-oligomerization was identified at the extreme amino terminus of Hip.
同寡聚体Hip蛋白在新合成多肽链的折叠以及已知与Hsc70和Hsp90相互作用的信号分子的构象调节中，与70 kDa热休克同源蛋白Hsc70协同作用。为了进一步评估Hip在蛋白质生物合成过程中的作用，我们启动了对Hip蛋白的结构-功能分析。通过使用酵母双杂交系统，Hip的Hsc70结合位点被定位到一个包含多个四肽重复序列和侧翼带电荷α螺旋的结构域。亲和层析证实了分离的Hip片段以及带有该区域的蛋白质融合体与Hsc70的ATP酶结构域以ATP和盐依赖的方式直接相互作用。Hip与ATP酶结构域的接触似乎主要由四肽重复序列之后带正电荷的α螺旋介导。此外，在Hip的极端氨基末端鉴定出了一个同寡聚化所需的结构域。
