Functional complementation of Hsp90 domains

Abstract

Hsp90s are essential ATP dependent molecular chaperones required for the maturation of numerous signal transduction proteins. They exist as obligatory homodimers consisting of an N‐, Middle, and C‐domain. In higher eukaryotes there are four hsp90 paralogs: two cytosolic isoforms, Hsp90α and Hsp90β, the ER paralog GRP94 and the mitochondrial paralog TRAP‐1. Despite the high sequence similarity between cytosolic Hsp90 and GRP94, structural and biochemical studies have elucidated different nucleotide induced conformational states which account for a 5–20 fold difference in ATPase rates. To further characterize the similarities and differences between Hsp90 and GRP94, we constructed chimeras between domains of cytosolic Hsp90 and GRP94 and tested the viability of these chimeras when expressed as the sole Hsp90 in yeast. We show for the first time that Grp94 domains can functionally complement yeast Hsp90 domains. In particular, chimeric constructs consisting of Grp94 N or Grp94 MC support growth in yeast while M domain substitutions are detrimental to cell growth. These results indicate the importance of ATP binding and highlight the MC interface as an important structural unit within hsp90s. These observations also suggest a conserved mechanism of action within the hsp90s family. Finally, the GRP N chimera may be used to develop paralog specific inhibitors. Funding: NIH 5F31GM086120, NIH 2R01CA095130