Crystal structure of yeast Sis1 peptide-binding fragment and Hsp70 Ssa1 C-terminal complex

Li, Jingzhi; Wu, Yunkun; Qian, Xiao Ming; Sha, Bingdong

doi:10.1042/bj20060618

Cited by 58 publications

(74 citation statements)

References 27 publications

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“…3b) by virtue of intramolecular hydrogen bonding between Pro-220 and Glu-223 of the Ssa1 peptide. The same Ssa1 peptide forms a ␤-strand in the structure of the unrelated Sis1 (Hsp40)-Ssa1 complex (31). Taken together, this shows that the C-terminal Ssa1 tail is versatile and can adopt different structures depending on its binding partner.…”

Section: Sec71-sec72 and Hsp70s In Post-translational Protein Translomentioning

confidence: 75%

Two alternative binding mechanisms connect the protein translocation Sec71-Sec72 complex with heat shock proteins

Tripathi

Mandon

Gilmore

et al. 2017

Journal of Biological Chemistry

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The biosynthesis of many eukaryotic proteins requires accurate targeting to and translocation across the endoplasmic reticulum membrane. Post-translational protein translocation in yeast requires both the Sec61 translocation channel, and a complex of four additional proteins: Sec63, Sec62, Sec71, and Sec72. The structure and function of these proteins are largely unknown. This pathway also requires the cytosolic Hsp70 protein Ssa1, but whether Ssa1 associates with the translocation machinery to target protein substrates to the membrane is unclear. Here, we use a combined structural and biochemical approach to explore the role of Sec71-Sec72 subcomplex in post-translational protein translocation. To this end, we report a crystal structure of the Sec71-Sec72 complex, which revealed that Sec72 contains a tetratricopeptide repeat (TPR) domain that is anchored to the endoplasmic reticulum membrane by Sec71. We also determined the crystal structure of this TPR domain with a C-terminal peptide derived from Ssa1, which suggests how Sec72 interacts with full-length Ssa1. Surprisingly, Ssb1, a cytoplasmic Hsp70 that binds ribosome-associated nascent polypeptide chains, also binds to the TPR domain of Sec72, even though it lacks the TPR-binding C-terminal residues of Ssa1. We demonstrate that Ssb1 binds through its ATPase domain to the TPR domain, an interaction that leads to inhibition of nucleotide exchange. Taken together, our results suggest that translocation substrates can be recruited to the Sec71-Sec72 complex either post-translationally through Ssa1 or cotranslationally through Ssb1.

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Section: Sec71-sec72 and Hsp70s In Post-translational Protein Translomentioning

confidence: 75%

Two alternative binding mechanisms connect the protein translocation Sec71-Sec72 complex with heat shock proteins

Tripathi

Mandon

Gilmore

et al. 2017

Journal of Biological Chemistry

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“…The basis for their differential function is due in part to each having a distinct chaperone module, which can specify the function of Sis1 or Ydj1 in chimeric molecules (19). We note, however, that Sis1 also has a more stable interaction with the Hsp70 C terminus than does Ydj1 (33). The C-terminal region of Hsp70 interacts with the peptide-binding groove in the chaperone module of Sis1, suggesting that there could be competitive interactions.…”

Section: Discussionmentioning

confidence: 95%

Ydj1 Protects Nascent Protein Kinases from Degradation and Controls the Rate of Their Maturation

Mandal

Nillegoda

Chen

et al. 2008

Molecular and Cellular Biology

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Ydj1 is a Saccharomyces cerevisiae Hsp40 molecular chaperone that functions with Hsp70 to promote polypeptide folding. We identified Ydj1 as being important for maintaining steady-state levels of protein kinases after screening several chaperones and cochaperones in gene deletion mutant strains. Pulse-chase analyses revealed that a portion of Tpk2 kinase was degraded shortly after synthesis in a ydj1⌬ mutant, while the remainder was capable of maturing but with reduced kinetics compared to the wild type. Cdc28 maturation was also delayed in the ydj1⌬ mutant strain. Ydj1 protects nascent kinases in different contexts, such as when Hsp90 is inhibited with geldanamycin or when CDC37 is mutated. The protective function of Ydj1 is due partly to its intrinsic chaperone function, but this is minor compared to the protective effect resulting from its interaction with Hsp70. SIS1, a type II Hsp40, was unable to suppress defects in kinase accumulation in the ydj1⌬ mutant, suggesting some specificity in Ydj1 chaperone action. However, analysis of chimeric proteins that contained the chaperone modules of Ydj1 or Sis1 indicated that Ydj1 promotes kinase accumulation independently of its client-binding specificity. Our results suggest that Ydj1 can both protect nascent chains against degradation and control the rate of kinase maturation.

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“…Our finding therefore suggests that the lid movement can allosterically regulate association/dissociation of the two domains. The C-terminal tail of the lid subdomain has been implicated in interactions of mammalian Hsp70s with DnaJ homologues (16,20,27,40,53). Allosteric regulation of the association of the ATPase domain and the SBD by the lid movement is therefore likely to play an important functional role in the operation of the DnaK-DnaJ chaperone machine.…”

Section: Resultsmentioning

confidence: 99%

Allosteric Coupling between the Lid and Interdomain Linker in DnaK Revealed by Inhibitor Binding Studies

Liebscher

Roujeinikova

2009

J Bacteriol

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The molecular chaperone DnaK assists protein folding and refolding, translocation across membranes, and regulation of the heat shock response. In Escherichia coli, the protein is a target for insect-derived antimicrobial peptides, pyrrhocoricins. We present here the X-ray crystallographic analysis of the E. coli DnaK substratebinding domain in complex with pyrrhocoricin-derived peptide inhibitors. The structures show that pyrrhocoricins act as site-specific, dual-mode (competitive and allosteric) inhibitors, occupying the substrate-binding tunnel and disrupting the latch between the lid and the ␤-sandwich. Our structural analysis revealed an allosteric coupling between the movements of the lid and the interdomain linker, identifying a previously unknown mechanism of the lid-mediated regulation of the chaperone cycle.DnaK is the bacterial molecular chaperone from the Hsp70 (70-kDa heat shock protein) family that assists many cellular processes involving proteins in their nonnative conformations, such as folding of newly synthesized proteins, protein translocation across membranes, refolding of misfolded and aggregated proteins, degradation of unstable proteins, and regulation of the heat shock response (for reviews, see references 10, 28, and 42). The chaperone function of DnaK is based on its ability to transiently bind to exposed stretches of hydrophobic residues in partially or fully unfolded proteins in an ATP-controlled fashion, thereby preventing aggregation and misfolding. DnaK recognizes short peptide sequences containing up to five consecutive hydrophobic residues (with leucine found frequently in the middle), flanked preferentially by basic residues (42, 43). The molecularchaperone activity is functionally linked with ATP hydrolysis; the substrate-binding and release cycle is driven by the switching between the ATP-bound state, with low affinity and a high exchange rate for substrates, and the ADP-bound state, with high affinity and a low exchange rate for substrates.In vivo, DnaK activity is supported by two cochaperones, GrpE, which facilitates the ADP/ATP exchange, and DnaJ, which stimulates ATP hydrolysis and thus aids the peptide capture (10,25,28,38,54). DnaJ itself also recognizes exposed stretches of hydrophobic residues in partially unfolded or denatured proteins, with specificity overlapping with that of DnaK (44). It is therefore thought that DnaJ serves as a scanning factor for DnaK by binding specific unfolded substrates and presenting them to the ATP-bound form of DnaK.Escherichia coli DnaK is composed of two domains: an Nterminal ATPase domain (residues 1 to 387) and a C-terminal substrate-binding domain (SBD) (residues 388 to 638) (10).The latter is made up of an 18-kDa ␤-sandwich subdomain that holds the substrate-binding cleft and a C-terminal ␣-helicalbundle "lid" subdomain that stabilizes the complex with the peptide substrate and controls the accessibility of the peptide binding site but does not interact with the substrate directly (5, 55). Removal of the lid subdomain by truncatio...

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Crystal structure of yeast Sis1 peptide-binding fragment and Hsp70 Ssa1 C-terminal complex

Cited by 58 publications

References 27 publications

Two alternative binding mechanisms connect the protein translocation Sec71-Sec72 complex with heat shock proteins

Two alternative binding mechanisms connect the protein translocation Sec71-Sec72 complex with heat shock proteins

Ydj1 Protects Nascent Protein Kinases from Degradation and Controls the Rate of Their Maturation

Allosteric Coupling between the Lid and Interdomain Linker in DnaK Revealed by Inhibitor Binding Studies

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