NMR structure determination of the Escherichia coli DnaJ molecular chaperone: secondary structure and backbone fold of the N-terminal region (residues 2-108) containing the highly conserved J domain.
DnaJ from Escherichia coli is a 376-amino acid protein that functions in conjunction with DnaK and GrpE as a chaperone machine. The N-terminal fragment of residues 2-108, DnaJ-(2-108), retains many of the activities of the full-length protein and contains a structural motif, the J domain of residues 2-72, which is highly conserved in a superfamily of proteins. In this paper, NMR spectroscopy was used to determine the secondary structure and the three-dimensional polypeptide backbone fold of DnaJ-(2-108). By using 13C/15N doubly labeled DnaJ-(2-108), nearly complete sequence-specific assignments were obtained for 1H, 15N, 13C alpha, and 13C beta, and about 40% of the peripheral aliphatic carbon resonances were also assigned. Four alpha-helices in polypeptide segments of residues 6-11, 18-31, 41-55, and 61-68 in the J domain were identified by sequential and medium-range nuclear Overhauser effects. For the J domain, the three-dimensional structure was calculated with the program DIANA from an input of 536 nuclear Overhauser effect upper-distance constraints and 52 spin-spin coupling constants. The polypeptide backbone fold is characterized by the formation of an antiparallel bundle of two long helices, residues 18-31 and 41-55, which is stabilized by a hydrophobic core of side chains that are highly conserved in homologous J domain sequences. The Gly/Phe-rich region from residues 77 to 108 is flexibly disordered in solution.
Institut für Molekularbiologie und Biophysik, Eidgenössische Technische Hochschule-Hönggerberg, Zürich, Switzerland.
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T
Szyperski
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Pellecchia
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Wall
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Georgopoulos
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Wüthrich
D000595:Amino Acid Sequence; D004926:Escherichia coli / Q000737:chemistry; D029968:Escherichia coli Proteins; D050956:HSP40 Heat-Shock Proteins; D006360:Heat-Shock Proteins / Q000648:ultrastructure*; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D018832:Molecular Chaperones / Q000648:ultrastructure*; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary
C494635:DnaJ protein, E coli; D029968:Escherichia coli Proteins; D050956:HSP40 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones