Structural analysis of the ribosome-associated complex (RAC) reveals an unusual Hsp70/Hsp40 interaction.
Yeast Zuotin and Ssz are members of the conserved Hsp40 and Hsp70 chaperone families, respectively, but compared with canonical homologs, they atypically form a stable heterodimer termed ribosome-associated complex (RAC). RAC acts as co-chaperone for another Hsp70 to assist de novo protein folding. In this study, we identified the molecular basis for the unusual Hsp70/Hsp40 pairing using amide hydrogen exchange (HX) coupled with mass spectrometry and mutational analysis. Association of Ssz with Zuotin strongly decreased the conformational dynamics mainly in the C-terminal domain of Ssz, whereas Zuotin acquired strong conformational stabilization in its N-terminal segment. Deletion of the highly flexible N terminus of Zuotin abolished stable association with Ssz in vitro and caused a phenotype resembling the loss of Ssz function in vivo. Thus, the C-terminal domain of Ssz, the N-terminal extension of Zuotin, and their mutual stabilization are the major structural determinants for RAC assembly. We furthermore found dynamic changes in the J-domain of Zuotin upon complex formation that might be crucial for RAC co-chaperone function. Taken together, we present a novel mechanism for converting Zuotin and Ssz chaperones into a functionally active dimer.
Fiaux
Jocelyne
J
Zentrum für Molekulare Biologie der Universität Heidelberg, DKFZ-ZMBH Alliance, INF282, 69120 Heidelberg, Germany.
Horst
Janina
J
Scior
Annika
A
Preissler
Steffen
S
Koplin
Ansgar
A
Bukau
Bernd
B
Deuerling
Elke
E
D004252:DNA Mutational Analysis; D019281:Dimerization; D004926:Escherichia coli / Q000378:metabolism; D050956:HSP40 Heat-Shock Proteins / Q000378:metabolism*; D018840:HSP70 Heat-Shock Proteins / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology; D013058:Mass Spectrometry; D018832:Molecular Chaperones / Q000737:chemistry / Q000502:physiology*; D010641:Phenotype; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D012270:Ribosomes / Q000737:chemistry*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000502:physiology*
D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins; D018832:Molecular Chaperones; C424445:SSZ1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C077019:ZUO1 protein, S cerevisiae