The Type I Hsp40 Zinc Finger-like Region Is Required for Hsp70 to Capture Non-native Polypeptides from Ydj1
Department of Cell and Developmental Biology, School of Medicine, University of North Carolina, Chapel Hill, North Carolina 27599-7090, USA. Journal of Biological Chemistry
(Impact Factor: 4.57).
02/2005; 280(1):695-702. DOI: 10.1074/jbc.M410645200
The cytosolic yeast Hsp40 Ydj1 contains a conserved zinc finger-like region (ZFLR), which has two zinc-binding domains (ZBD), that helps regulate and specify Hsp70 function. To investigate the mechanism for Ydj1 ZFLR action, ZBDI and ZBDII mutants were constructed and characterized. ZBDII mutants exhibited temperature-sensitive growth defects, but yeast tolerated mutation of ZBDI. However, ZBDI and ZBDII mutants were defective at facilitating androgen receptor (AR) folding. Defective AR folding was associated with the accumulation of complexes between AR and Ydj1 ZFLR mutants and a reduction in Hsp70.AR complex formation. Purified Ydj1 ZBDI and ZBDII mutants could bind non-native polypeptides but could not deliver luciferase to Hsp70 and were defective at luciferase refolding. Interestingly, the ability of Ydj1 to synergize with Hsp70 to suppress thermally induced protein aggregation was blocked by mutation of ZBDII, but not ZBDI. Hence, ZBDII is required for yeast to survive heat stress because it is essential for Ydj1 to cooperate with Hsp70 to suppress protein aggregation. On the other hand, protein folding is dependent upon the action of both ZBDI and ZBDII because each is required for Hsp70 to capture non-native polypeptides from Ydj1.
Available from: Hyun Young Yu
- "roles in their function   . We constructed chimeric genes substituting the Zn binding domains of Ydj1 and Xdj1 for that of DnaJA2, generating DnaJA2 Zn-Ydj1 and DnaJA2 Zn-Xdj1 respectively. "
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ABSTRACT: At their C-termini, cytosolic Hsp70s have an EEVD tetrapeptide that interacts with J-protein co-chaperones of the B, but not A, class. This interaction is required for partnering with yeast B-type J-proteins in protein folding. Here we report conservation of this feature. Human B-type J-proteins also have a stringent EEVD requirement. Human A-type J-proteins function less well than their yeast orthologs with Hsp70ΔEEVD. Changes in the zinc binding domain, a domain absent in B-type J-proteins, overcomes this partial EEVD dependence. Our results suggest that the structurally similar A- and B-class J-proteins of the cytosol have evolved conserved, yet distinct, features that enhance specialized functionality of Hsp70 machinery.
Copyright © 2015. Published by Elsevier B.V.
FEBS letters 08/2015; 589(19 Pt B). DOI:10.1016/j.febslet.2015.07.040 · 3.17 Impact Factor
Available from: Yusuf Tutar
- "Hsp40 family consists of more than 100 members and some Hsp40s themselves show ATPase activity. These Hsp40s contribute to substrate protein folding as evidenced by luciferase folding experiments      . "
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ABSTRACT: Heat shock proteins (Hsps) are highly conserved proteins and have cytoprotective role for maintaining cellular protein conformation. Hsps not only keep proteins in their native state but also involve in several essential biochemical process. This review summarizes structural properties of Hsps (Hsp70, Hsp40, Hsp90, Hsp100, Hsp60, sHsps, and Nucleotide Exchange Factors) and explains their roles in aging, apoptosis, cancer, neurodegeneration, cardio-vascular diseases, obesity and diabetes mellitus, and housekeeping.
Frontiers in Protein and Peptide Sciences Volume 1, 1 edited by Ben Dunn, 07/2014: chapter Heat Shock Response Agents and the Diseases: pages 139-160 (22); Bentham., ISBN: 978-1-60805-863-1
Available from: Alexander Kai Buell
- "The J domain of Ydj1p, which can stimulate the ATPase activity of Ssa1p, is important for curing of [URE3] in vivo; hence, it has been suggested that the curing effect of Ydj1p on [URE3] is indirect and occurs via its effect on Ssa1p [79,80]. We therefore tested the Ydj1p mutant H34Q, which is deficient in stimulating the ATPase activity of HSP70 [81,82] and also disables curing of [URE3] by Ydj1p [79,80]. We found that Ydj1p H34Q shows significantly weaker inhibition of Ure2p fibrillation in vitro compared with WT Ydj1p, both in the presence (figure 3b) and absence (figure 3c) of Ssa1p, which could account for the weaker curing effect of Ydj1p H34Q in vivo [79,80]. "
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ABSTRACT: Ure2p is the protein determinant of the Saccharomyces cerevisiae prion state [URE3]. Constitutive overexpression of the HSP70 family member SSA1 cures cells of [URE3]. Here, we show that Ssa1p increases the lag time of Ure2p fibril formation in vitro in the presence or absence of nucleotide. The presence of the HSP40 co-chaperone Ydj1p has an additive effect on the inhibition of Ure2p fibril formation, whereas the Ydj1p H34Q mutant shows reduced inhibition alone and in combination with Ssa1p. In order to investigate the structural basis of these effects, we constructed and tested an Ssa1p mutant lacking the ATPase domain, as well as a series of C-terminal truncation mutants. The results indicate that Ssa1p can bind to Ure2p and delay fibril formation even in the absence of the ATPase domain, but interaction of Ure2p with the substrate-binding domain is strongly influenced by the C-terminal lid region. Dynamic light scattering, quartz crystal microbalance assays, pull-down assays and kinetic analysis indicate that Ssa1p interacts with both native Ure2p and fibril seeds, and reduces the rate of Ure2p fibril elongation in a concentration-dependent manner. These results provide new insights into the structural and mechanistic basis for inhibition of Ure2p fibril formation by Ssa1p and Ydj1p.
Philosophical Transactions of The Royal Society B Biological Sciences 05/2013; 368(1617):20110410. DOI:10.1098/rstb.2011.0410 · 7.06 Impact Factor
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