Group II archaeal chaperonin recognition of partially folded human γD-crystallin mutants.

The features in partially folded intermediates that allow the group II chaperonins to distinguish partially folded from native states remain unclear. The archaeal group II chaperonin from Methanococcus Mauripaludis (Mm-Cpn) assists the in vitro refolding of the well-characterized -sheet lens protein human D-crystallin (HD-Crys). The domain interface and buried cores of this Greek key conformation include side chains, which might be exposed in partially folded intermediates. We sought to assess whether particular features buried in the native state, but absent from the native protein surface, might serve as recognition signals. The features tested were (a) paired aromatic side chains, (b) side chains in the interface between the duplicated domains of HD-Crys, and (c) side chains in the buried core which result in congenital cataract when substituted. We tested the Mm-Cpn suppression of aggregation of these HD-Crys mutants upon dilution out of denaturant. Mm-Cpn was capable of suppressing the off-pathway aggregation of the three classes of mutants indicating that the buried residues were not recognition signals. In fact, Mm-Cpn recognized the HD-Crys mutants better than (wild-type) WT and refolded most mutant HD-Crys to levels twice that of WT HD-Crys. This presumably represents the increased population or longer lifetimes of the partially folded intermediates of the mutant proteins. The results suggest that Mm-Cpn does not recognize the features of HD-Crys testedpaired aromatics, exposed domain interface, or destabilized corebut rather recognizes other features of the partially folded -sheet conformation that are absent or inaccessible in the native state of HD-Crys.