HSP 90 Assoc. with Steroid Receptors & Other Proteins

描述(申请人提供)：HSP90是细胞内关键的分子伴侣蛋白。它参与产生和维持许多重要的调节蛋白的活性状态，包括一些转录因子和蛋白激酶。类固醇受体的活性依赖于HSP90的伴侣作用，我们已经使用孕激素受体(PR)作为模型客户蛋白来研究HSP90的作用机制。这些研究的目的是确定HSP90与客户蛋白结合的性质和机制，并获得关于这一重要相互作用的结果的信息。提出了三个具体目标。在目标1中，将开发用于直接描述HSP90与PR结合的方法。这种相互作用是复杂的，通常需要额外的共同监护人。然而，通过温和的加热，一个人可以诱导HSP90与PR结合，而不需要其他蛋白质的帮助，从而允许在简化的系统中研究客户结合的几个方面。在目标2中，将研究HSP90与小肽的结合，作为了解HSP90客户结合活性的第二个途径。将使用噬菌体展示方法从随机文库中选择多肽，并将鉴定它们在HSP90上的结合部位。在目标3中，关于HSP90的这一新信息将被用于研究客户结合在更复杂的系统中的作用和调节，在该系统中，HSP90与其他伴侣和辅助伴侣合作产生活性形式的客户蛋白。目前，HSP90选择和结合客户蛋白的机制尚不清楚。这些研究将对这一机制提供有价值的新见解，这是理解HSP90生物学意义的关键。

DESCRIPTION (provided by applicant): hsp90 is a key molecular chaperone protein in the cell. It participates in producing and maintaining the active states of many important regulatory proteins including a number of transcription factors and protein kinases. The activities of steroid receptors are dependent on hsp90 chaperoning and we have used the progesterone receptor (PR) as a model client protein for studies on the mechanism of hsp90 action. The objective of the studies proposed is to define the properties and the mechanism of hsp90 binding to client proteins and to gain information on the consequences of this important interaction. Three specific aims are proposed. In Aim 1, methods will be developed to use in describing directly the binding of hsp90 to PR. This interaction is complex and normally requires additional co-chaperones. However, by mild heating one can induce hsp90 to bind PR without assistance by other proteins allowing the study of several aspects of client binding in a simplified system. In Aim 2, the binding of hsp90 to small peptides will be studied as a second approach to understanding the client binding activity of hsp90. Peptides will be selected from random libraries using phage display methods and their binding sites on hsp90 will be characterized. In Aim 3, this new information on hsp90 will be used to study the roles and the regulation of client binding in the more complex system where hsp90 cooperates with other chaperones and co-chaperones to produce active forms of client proteins. At present, the mechanism by which hsp90 selects and binds client proteins is unknown. These studies should provide valuable new insights into this mechanism which is key to understanding the biological significance of hsp90.