Fast Closure of N‑Terminal Long Loops but Slow
Formation of β Strands Precedes the Folding Transition State
of <i>Escherichia coli</i> Adenylate Kinase
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Abstract
The
nature of the earliest steps of the initiation of the folding
pathway of globular proteins is still controversial. To elucidate
the role of early closure of long loop structures in the folding transition,
we studied the folding kinetics of subdomain structures in <i>Escherichia coli</i> adenylate kinase (AK) using Förster
type resonance excitation energy transfer (FRET)-based methods. The
overall folding rate of the AK molecule and of several segments that
form native β strands is 0.5 ± 0.3 s<sup>–1</sup>, in sharp contrast to the 1000-fold faster closure of three long
loop structures in the CORE domain. A FRET-based “double kinetics”
analysis revealed complex transient changes in the initially closed
N-terminal loop structure that then opens and closes again at the
end of the folding pathway. The study of subdomain folding <i>in situ</i> suggests a hierarchic ordered folding mechanism,
in which early and rapid cross-linking by hydrophobic loop closure
provides structural stabilization at the initiation of the folding
pathway