The crystal structure of the yeast Hsp40 Ydj1 complexed with its peptide substrate.

The mechanisms by which Hsp40 functions as a molecular chaperone to recognize and bind non-native polypeptides is not understood. We have identified a peptide substrate for Ydj1, a member of the type I Hsp40 from yeast. The structure of the Ydj1 peptide binding fragment and its peptide substrate complex was determined to 2.7 A resolution. The complex structure reveals that Ydj1 peptide binding fragment forms an L-shaped molecule constituted by three domains. The domain I exhibits a similar protein folds as domain III while the domain II contains two Zinc finger motifs. The peptide substrate binds Ydj1 by forming an extra beta strand with domain I of Ydj1. The Leucine residue in the middle of the peptide substrate GWLYEIS inserts its side chain into a hydrophobic pocket formed on the molecular surface of Ydj1 domain I. The Zinc finger motifs located in the Ydj1 domain II are not in the vicinity of peptide substrate binding site.

[1]  Z. Otwinowski,et al.  [20] Processing of X-ray diffraction data collected in oscillation mode. , 1997, Methods in enzymology.

[2]  Soojin Lee,et al.  Identification of Essential Residues in the Type II Hsp40 Sis1 That Function in Polypeptide Binding* , 2002, The Journal of Biological Chemistry.

[3]  A. Caplan,et al.  Characterization of YDJ1: a yeast homologue of the bacterial dnaJ protein , 1991, The Journal of cell biology.

[4]  Craig M. Ogata,et al.  Structural Analysis of Substrate Binding by the Molecular Chaperone DnaK , 1996, Science.

[5]  F. Hartl,et al.  A zinc finger‐like domain of the molecular chaperone DnaJ is involved in binding to denatured protein substrates. , 1996, The EMBO journal.

[6]  P. Christen,et al.  Kinetics of molecular chaperone action. , 1994, Science.

[7]  D. Cyr,et al.  Protein Folding Activity of Hsp70 Is Modified Differentially by the Hsp40 Co-chaperones Sis1 and Ydj1* , 1998, The Journal of Biological Chemistry.

[8]  Bernd Bukau,et al.  Its substrate specificity characterizes the DnaJ co‐chaperone as a scanning factor for the DnaK chaperone , 2001, The EMBO journal.

[9]  C. Georgopoulos,et al.  The NH2-terminal 108 amino acids of the Escherichia coli DnaJ protein stimulate the ATPase activity of DnaK and are sufficient for lambda replication. , 1994, The Journal of biological chemistry.

[10]  F. Hartl,et al.  Successive action of DnaK, DnaJ and GroEL along the pathway of chaperone-mediated protein folding , 1992, Nature.

[11]  J. Reinstein,et al.  Mechanism of regulation of hsp70 chaperones by DnaJ cochaperones. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[12]  J. Zou,et al.  Improved methods for building protein models in electron density maps and the location of errors in these models. , 1991, Acta crystallographica. Section A, Foundations of crystallography.

[13]  K. Sharp,et al.  Protein folding and association: Insights from the interfacial and thermodynamic properties of hydrocarbons , 1991, Proteins.

[14]  J. Rothman,et al.  Peptide-binding specificity of the molecular chaperone BiP , 1991, Nature.

[15]  W. Hendrickson.  Determination of macromolecular structures from anomalous diffraction of synchrotron radiation. , 1991, Science.

[16]  B. Matthews.  Solvent content of protein crystals. , 1968, Journal of molecular biology.

[17]  E. Craig,et al.  An Essential Role for the Substrate-Binding Region of Hsp40s in Saccharomyces cerevisiae , 2001, The Journal of cell biology.

[18]  R J Read,et al.  Crystallography & NMR system: A new software suite for macromolecular structure determination. , 1998, Acta crystallographica. Section D, Biological crystallography.

[19]  Mike Carson,et al.  Ribbon models of macromolecules , 1987 .

[20]  P. Sigler,et al.  The Crystal Structure of a GroEL/Peptide Complex Plasticity as a Basis for Substrate Diversity , 1999, Cell.

[21]  F. Hartl.  Molecular chaperones in cellular protein folding , 1996, Nature.

[22]  B. Sha,et al.  Direct interactions between molecular chaperones heat-shock protein (Hsp) 70 and Hsp40: yeast Hsp70 Ssa1 binds the extreme C-terminal region of yeast Hsp40 Sis1. , 2002, The Biochemical journal.

[23]  Bernd Bukau,et al.  The Hsp70 and Hsp60 Chaperone Machines , 1998, Cell.

[24]  T. Rapoport,et al.  J proteins catalytically activate Hsp70 molecules to trap a wide range of peptide sequences. , 1998, Molecular cell.

[25]  J. Scott,et al.  Searching for peptide ligands with an epitope library. , 1990, Science.

[26]  C. Georgopoulos,et al.  Structure-Function Analysis of the Zinc Finger Region of the DnaJ Molecular Chaperone* , 1996, The Journal of Biological Chemistry.

[27]  P E Wright,et al.  Solution structure of the cysteine-rich domain of the Escherichia coli chaperone protein DnaJ. , 2000, Journal of molecular biology.

[28]  G. Waksman,et al.  Chaperone Priming of Pilus Subunits Facilitates a Topological Transition that Drives Fiber Formation , 2002, Cell.

[29]  D. Cyr,et al.  The crystal structure of the peptide-binding fragment from the yeast Hsp40 protein Sis1. , 2000, Structure.

[30]  F. Hartl,et al.  Molecular Chaperones in the Cytosol: from Nascent Chain to Folded Protein , 2002, Science.