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Cosolvent-induced transformation of a death domain tertiary structure.
- Source :
-
Proceedings of the National Academy of Sciences of the United States of America [Proc Natl Acad Sci U S A] 2002 Aug 20; Vol. 99 (17), pp. 11151-6. Date of Electronic Publication: 2002 Aug 12. - Publication Year :
- 2002
-
Abstract
- The death domain (DD) of the protein kinase Pelle adopts a six-helix bundle fold in the crystal structure of the complex with its dimerization partner, Tube-DD. However, in crystals obtained from a solution of 45% 2-methyl-2,4-pentanediol (MPD), the C-terminal half of Pelle-DD folds into a single helix, and the N-terminal half of the molecule is disordered. The helical segment forms an antiparallel dimer with the corresponding helix of a symmetry-related molecule, and together they form extensive lattice interactions similar in number, composition, and buried surface to those in the six-helix bundle of the native fold. Secondary structure analysis by heteronuclear nuclear magnetic resonance spectroscopy (NMR) demonstrates that Pelle-DD adopts a six-helix bundle fold in aqueous solution. The fold is perturbed by MPD, with the largest chemical shift changes in one helix and two loop regions that encompass the Tube-DD binding site. Pelle-DD is stable to urea denaturation with a folding free energy of 7.9 kcal/mol at 25 degrees C but is destabilized, with loss of urea binding sites, in the presence of MPD. The data are consistent with a cosolvent denaturation model in which MPD denatures the N terminus of Pelle-DD but induces the C terminus to form a more compact structure and aggregate. A similar perturbation in vivo might occur at the plasma membrane and could have consequences for Pelle-mediated regulation. Generally, crystallographers should be aware that high concentrations of MPD or related cosolvents can alter the tertiary structure of susceptible proteins.
- Subjects :
- Amino Acid Sequence
Animals
Binding Sites
Crystallography, X-Ray methods
Drosophila melanogaster enzymology
Enzyme Stability
Kinetics
Models, Molecular
Molecular Sequence Data
Protein Denaturation
Protein Structure, Tertiary
Recombinant Proteins chemistry
Recombinant Proteins metabolism
Solvents
Urea
Drosophila Proteins
Protein Serine-Threonine Kinases chemistry
Protein Serine-Threonine Kinases metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 0027-8424
- Volume :
- 99
- Issue :
- 17
- Database :
- MEDLINE
- Journal :
- Proceedings of the National Academy of Sciences of the United States of America
- Publication Type :
- Academic Journal
- Accession number :
- 12177432
- Full Text :
- https://doi.org/10.1073/pnas.172188399