Collagen processing and molecular chaperones : molecular anatomy based on the domain structures

胶原蛋白分子的翻译后加工已被研究与分子伴侣的功能有关。我们首先研究了Hsp47（一种胶原特异性分子伴侣）在疤痕形成中的作用，即损伤引发的过多和异常的胶原合成。我们发现Hsp47和I型胶原蛋白在胎鼠伤口中没有被诱导，而在新生大鼠伤口中却被增强。因此，我们试图通过抗治疗治疗来抑制胶原沉积。结果表明，抗Hsp47的反义寡核苷酸在体外（原代培养成纤维细胞）和体内（背创面）有效地阻止了新生大鼠损伤后瘢痕的形成。这些发现有力地表明，Hsp47可能是预防腭裂等外科手术中瘢痕形成的潜在靶点。我们还研究了Hsp90的作用机制。大肠杆菌HtpG是一种与哺乳动物Hsp90同源的细菌，与人类Hsp90在一级结构水平上有更多的自由结构域。N端、中端和C端结构域分别称为N、M和C结构域。Hsp90的客户端结合活性主要定位于N结构域。结构域之间存在两种相互作用：N和M结构域之间的分子内相互作用；以及M和C结构域之间的分子间相互作用。后者的相互作用介导了HSP90二聚体的形成。前者相互作用的释放伴随着高温诱导的Hsp90分子伴侣的客户端结合活性的激活。即位于N结构域的客户端结合位点被M结构域掩盖，但热休克破坏了相互作用。异常酵母HSP90 （Hsc82）的表达证实了N和M结构域相互作用的重要性，其中M结构域不能与N结构域相互作用。因此，我们提出释放分子内相互作用作为热诱导激活Hsp90分子伴侣的机制。少

Post-translational processing of collagen molecules has been investigated in relation to the function of molecular chaperones. We first investigated the role of Hsp47, a collagen specific molecular chaperone, in scar formation, i.e. excessive and aberrant collagen synthesis triggered by the wounding. We found that Hsp47 as well as type I collagen was not induced in fetal rat wound, in contrast to their enhancement in neonatal rat wound. Hence, we tried to suppress collagen deposition by anti-therapeutic treatment. As a result, scar formation after wounding of neonatal rats was efficiently prevented by an anti-sense oligonucleotide against Hsp47 in vitro (primary-cultured fibroblasts) and in vivo (back wound). These findings strongly suggested that Hsp47 could be a potential target for prevention of scar formation on surgical operations, such as cleft palate.We also investigated the functional mechanism of Hsp90. E. coli HtpG, a bacterial homologue of mammalian Hsp90, was composed of th … More ree domains at the primary structure level as those of human Hsp90. The N-terminal, middle and C-terminal domains were referred to N, M and C domains, respectively. The client-binding activity of Hsp90 was primarily localized in N domain. There were two interactions between the domains : an intramolecular interaction between N and M domains ; and an intermolecular interaction between M and C domains. The latter interaction mediated dimer formation of HSP90. Liberation of the former interaction accompanied the high temperature-induced activation of the client-binding activity of Hsp90 molecular chaperone. That is, the client-binding site located in N domain was concealed by M domain, but heat shock disrupted the interaction. Importance of the interaction between N and M domains was confirmed by expression of aberrant yeast HSP90 (Hsc82), of which M domain could not interact with N domain. We therefore propose the liberation of the intra-molecular interaction as the mechanism of heat-induced activation of Hsp90 molecular chaperone. Less