Unfolding domains of recombinant fusion alpha alpha-tropomyosin.
The thermal unfolding of the coiled-coil alpha-helix of recombinant alpha alpha-tropomyosin from rat striated muscle containing an additional 80-residue peptide of influenza virus NS1 protein at the N-terminus (fusion-tropomyosin) was studied with circular dichroism and fluorescence techniques. Fusion-tropomyosin unfolded in four cooperative transitions: (1) a pretransition starting at 35 degrees C involving the middle of the molecule; (2) a major transition at 46 degrees C involving no more than 36% of the helix from the C-terminus; (3) a major transition at 56 degrees C involving about 46% of the helix from the N-terminus; and (4) a transition from the nonhelical fusion domain at about 70 degrees C. Rabbit skeletal muscle tropomyosin, which lacks the fusion peptide but has the same tropomyosin sequence, does not exhibit the 56 degrees C or 70 degrees C transition. The very stable fusion unfolding domain of fusion-tropomyosin, which appears in electron micrographs as a globular structural domain at one end of the tropomyosin rod, acts as a cross-link to stabilize the adjacent N-terminal domain. The least stable middle of the molecule, when unfolded, acts as a boundary to allow the independent unfolding of the C-terminal domain at 46 degrees C from the stabilized N-terminal unfolding domain at 56 degrees C. Thus, strong localized interchain interactions in coiled-coil molecules can increase the stability of neighboring domains.
Ishii
Y
Y
Department of Muscle Research, Boston Biomedical Research Institute, Massachusetts 02114.
Hitchcock-DeGregori
S
S
Mabuchi
K
K
Lehrer
S S
SS
D000818:Animals; D009980:Influenza A virus / Q000235:genetics; D017510:Protein Folding; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000737:chemistry*; D013696:Temperature; D014335:Tropomyosin / Q000737:chemistry* / Q000235:genetics; D017361:Viral Nonstructural Proteins / Q000737:chemistry / Q000235:genetics
D011993:Recombinant Fusion Proteins; D014335:Tropomyosin; D017361:Viral Nonstructural Proteins