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  The ATP-ase cycle of the endoplasmic chaperone GRP94

Frey, S., Leskovar, A., Reinstein, J., & Buchner, J. (2007). The ATP-ase cycle of the endoplasmic chaperone GRP94. The Journal of Biological Chemistry, 282(49), 35612-35620. doi:10.1074/jbc.M704647200.

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Item Permalink: http://hdl.handle.net/11858/00-001M-0000-002C-2014-7 Version Permalink: http://hdl.handle.net/11858/00-001M-0000-002C-2015-5
Genre: Journal Article
Alternative Title : The ATP-ase cycle of the endoplasmic chaperone GRP94

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JBiolChem_282_2007_35612.pdf (Any fulltext), 309KB
 
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 Creators:
Frey, Stephan, Author
Leskovar, Adriane1, Author              
Reinstein, Jochen1, Author              
Buchner, Johannes, Author
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1Department of Biomolecular Mechanisms, Max Planck Institute for Medical Research, Max Planck Society, ou_1497700              

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 Abstract: Grp94, the Hsp90 paralog of the endoplasmic reticulum, plays a crucial role in protein secretion. Like cytoplasmic Hsp90, Grp94 is regulated by nucleotide binding to its N-terminal domain. However, the question of whether Grp94 hydrolyzes ATP was controversial. This sets Grp94 apart from other members of the Hsp90 family where a slow but specific turnover of ATP has been unambiguously established. In this study we aimed at analyzing the nucleotide binding properties and the potential ATPase activity of Grp94. We show here that Grp94 has an ATPase activity comparable with that of yeast Hsp90 with a k(cat) of 0.36 min(-1) at 25 degrees C. Kinetic and equilibrium constants of the partial reactions of the ATPase cycle were determined using transient kinetic methods. Nucleotide binding appears to be tighter compared with other Hsp90s investigated, with dissociation constants (K(D)) of approximately 4 microm for ADP, ATP, and AMP-PCP. Interestingly, all nucleotides and inhibitors (radicicol, 5'-N-ethylcarboxamidoadenosine) studied here bind with similar rate constants for association (0.2-0.3 x 10(6) M(-1) s(-1)). Furthermore, there is a marked difference from cytosolic Hsp90s in that after binding, the ATP molecule does not seem to become trapped by conformational changes in Grp94. Grp94 stays predominantly in the open state concerning the nucleotide-binding pocket as evidenced by kinetic analyses. Thus, Grp94 shows mechanistically important differences in the interaction with adenosine nucleotides, but the basic hydrolysis reaction seems to be conserved between cytosolic and endoplasmic members of the Hsp90 family.

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Language(s): eng - English
 Dates: 2007-09-142007-06-062007-10-092007-10-092007-12-07
 Publication Status: Published in print
 Pages: 9
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 Rev. Type: Peer
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Title: The Journal of Biological Chemistry
  Other : JBC
Source Genre: Journal
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Publ. Info: Baltimore, etc. : American Society for Biochemistry and Molecular Biology [etc.]
Pages: - Volume / Issue: 282 (49) Sequence Number: - Start / End Page: 35612 - 35620 Identifier: ISSN: 0021-9258
CoNE: https://pure.mpg.de/cone/journals/resource/954925410826_1