Thioredoxin fold as homodimerization module in the putative chaperone ERp29: NMR structures of the domains and experimental model of the 51 kDa dimer.
BACKGROUND
ERp29 is a ubiquitously expressed rat endoplasmic reticulum (ER) protein conserved in mammalian species. Fold predictions suggest the presence of a thioredoxin-like domain homologous to the a domain of human protein disulfide isomerase (PDI) and a helical domain similar to the C-terminal domain of P5-like PDIs. As ERp29 lacks the double-cysteine motif essential for PDI redox activity, it is suggested to play a role in protein maturation and/or secretion related to the chaperone function of PDI. ERp29 self-associates into 51 kDa dimers and also higher oligomers.


RESULTS
3D structures of the N- and C-terminal domains determined by NMR spectroscopy confirmed the thioredoxin fold for the N-terminal domain and yielded a novel all-helical fold for the C-terminal domain. Studies of the full-length protein revealed a short, flexible linker between the two domains, homodimerization by the N-terminal domain, and the presence of interaction sites for the formation of higher molecular weight oligomers. A gadolinium-based relaxation agent is shown to present a sensitive tool for the identification of macromolecular interfaces by NMR.


CONCLUSIONS
ERp29 is the first eukaryotic PDI-related protein for which the structures of all domains have been determined. Furthermore, an experimental model of the full-length protein and its association states was established. It is the first example of a protein where the thioredoxin fold was found to act as a specific homodimerization module, without covalent linkages or supporting interactions by further domains. A homodimerization module similar as in ERp29 may also be present in homodimeric human PDI.
Liepinsh
E
E
Department of Medical Biochemistry and Biophysics, Karolinska Institute, 171 77, Stockholm, Sweden.
Baryshev
M
M
Sharipo
A
A
Ingelman-Sundberg
M
M
Otting
G
G
Mkrtchian
S
S
D000595:Amino Acid Sequence; D019281:Dimerization; D004721:Endoplasmic Reticulum / Q000737:chemistry*; D006360:Heat-Shock Proteins / Q000737:chemistry*; D008958:Models, Molecular; D018832:Molecular Chaperones / Q000737:chemistry*; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D011487:Protein Conformation; D017510:Protein Folding; D017386:Sequence Homology, Amino Acid; D013879:Thioredoxins / Q000737:chemistry*
C491738:ERP29 protein, human; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones; D013879:Thioredoxins