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A distinct 14 residue site triggers coiled‐coil formation in cortexillin I
Author(s) -
Steinmetz Michel O.,
Stock Alexander,
Schulthess Therese,
Landwehr Ruth,
Lustig Ariel,
Faix Jan,
Gerisch Günther,
Aebi Ueli,
Kammerer Richard A.
Publication year - 1998
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1093/emboj/17.7.1883
Subject(s) - coiled coil , heptad repeat , circular dichroism , crystallography , trimer , protein folding , peptide , helix (gastropod) , folding (dsp implementation) , biophysics , protein structure , dimer , biology , peptide sequence , chemistry , biochemistry , ecology , organic chemistry , engineering , snail , electrical engineering , gene
We have investigated the process of the assembly of the Dictyostelium discoideum cortexillin I oligomerization domain (Ir) into a tightly packed, two‐stranded, parallel coiled‐coil structure using a variety of recombinant polypeptide chain fragments. The structures of these Ir fragments were analyzed by circular dichroism spectroscopy, analytical ultracentrifugation and electron microscopy. Deletion mapping identified a distinct 14 residue site within the Ir coiled coil, Arg311–Asp324, which was absolutely necessary for dimer formation, indicating that heptad repeats alone are not sufficient for stable coiled‐coil formation. Moreover, deletion of the six N‐terminal heptad repeats of Ir led to the formation of a four‐ rather than a two‐helix structure, suggesting that the full‐length cortexillin I coiled‐coil domain behaves as a cooperative folding unit. Most interestingly, a 16 residue peptide containing the distinct coiled‐coil ‘trigger’ site Arg311–Asp324 yielded ∼30% helix formation as monomer, in aqueous solution. pH titration and NaCl screening experiments revealed that the peptide's helicity depends strongly on pH and ionic strength, indicating that electrostatic interactions by charged side chains within the peptide are critical in stabilizing its monomer helix. Taken together, these findings demonstrate that Arg311–Asp324 behaves as an autonomous helical folding unit and that this distinct Ir segment controls the process of coiled‐coil formation of cortexillin I.