Prying open single GroES ring complexes by force reveals cooperativity across domains.

ABSTRACT: Understanding how the mechanical properties of a protein complex emerge from the interplay of intra- and interchain interactions is vital at both fundamental and applied levels. To investigate whether interdomain cooperativity affects protein mechanical strength, we employed single-molecule force spectroscopy to probe the mechanical stability of GroES, a homoheptamer with a domelike quaternary stucture stabilized by intersubunit interactions between the first and last ?-strands of adjacent domains. A GroES variant was constructed in which each subunit of the GroES heptamer is covalently linked to adjacent subunits by tripeptide linkers and folded domains of protein L are introduced to the heptamer's termini as handle molecules. The force-distance profiles for GroES unfolding showed, for the first time that we know of, a mechanical phenotype whereby seven distinct force peaks, with alternating behavior of unfolding force and contour length (?L(c)), were observed with increasing unfolding-event number. Unfolding of (GroES)(7) is initiated by breakage of the interface between domains 1 and 7 at low force, which imparts a polarity to (GroES)(7) that results in two distinct mechanical phenotypes of these otherwise identical protein domains. Unfolding then proceeds by peeling domains off the domelike native structure by sequential repetition of the denaturation of mechanically weak (unfoldon 1) and strong (unfoldon 2) units. These results indicate that domain-domain interactions help to determine the overall mechanical strength and unfolding pathway of the oligomeric structure. These data reveal an unexpected richness in the mechanical behavior of this homopolyprotein, yielding a complex with greater mechanical strength and properties distinct from those that would be apparent for GroES domains in isolation.

SUBMITTER: Ikeda-Kobayashi A 

PROVIDER: S-EPMC3328714 | biostudies-literature | 2012 Apr

REPOSITORIES: biostudies-literature