INVESTIGADORES
FERREIRO Diego Ulises
artículos
Título:
Stabilizing IkappaBalpha by "consensus" design.
Autor/es:
FERREIRO DU, CERVANTES CF, TRUHLAR SM, CHO SS, WOLYNES PG, KOMIVES EA.
Revista:
JOURNAL OF MOLECULAR BIOLOGY
Referencias:
Año: 2007 vol. 365 p. 1201 - 1216
ISSN:
0022-2836
Resumen:
IkappaBalpha is the major regulator of
transcription factor NF-kappaB function. The ankyrin repeat region of
IkappaBalpha mediates specific interactions with NF-kappaB dimers, but
ankyrin repeats 1, 5 and 6 display a highly dynamic character when not
in complex with NF-kappaB. Using chemical denaturation, we show here
that IkappaBalpha displays two folding transitions: a non-cooperative
conversion under weak perturbation, and a major cooperative folding
phase upon stronger insult. Taking advantage of a native Trp residue in
ankyrin repeat (AR) 6 and engineered Trp residues in AR2, AR4 and AR5,
we show that the cooperative transition involves AR2 and AR3, while the
non-cooperative transition involves AR5 and AR6. The major structural
transition can be affected by single amino acid substitutions
converging to the "consensus" ankyrin repeat sequence, increasing the
native state stability significantly. We further characterized the
structural and dynamic properties of the native state ensemble of
IkappaBalpha and the stabilized mutants by H/(2)H exchange mass
spectrometry and NMR. The solution experiments were complemented with
molecular dynamics simulations to elucidate the microscopic origins of
the stabilizing effect of the consensus substitutions, which can be
traced to the fast conformational dynamics of the folded ensemble.