Folding Mechanism and Aggregation Propensity of the KH0 Domain of FMRP and Its R138Q Pathological Variant

The K-homology (KH) domains are small, structurally conserved domains found in proteins of different origins characterized by a central conserved beta alpha alpha beta core and a GxxG motif in the loop between the two helices of the KH core. In the eukaryotic KHI type, additional alpha beta elements decorate the core at the C-terminus. Proteins containing KH domains perform different functions and several diseases have been associated with mutations in these domains, including those in the fragile X mental retardation protein (FMRP). FMRP is an RNA-binding protein crucial for the control of RNA metabolism whose lack or mutations lead to fragile X syndrome (FXS). Among missense mutations, the R138Q substitution is in the KH0 degenerated domain lacking the classical GxxG motif. By combining equilibrium and kinetic experiments, we present a characterization of the folding mechanism of the KH0 domain from the FMRP wild-type and of the R138Q variant showing that in both cases the folding mechanism implies the accumulation of an on-pathway transient intermediate. Moreover, by exploiting a battery of biophysical techniques, we show that the KH0 domain has the propensity to form amyloid-like aggregates in mild conditions in vitro and that the R138Q mutation leads to a general destabilization of the protein and to an increased fibrillogenesis propensity.

Automated summary

K-homology (KH) domains are structurally conserved domains found in proteins of different origins, which play important roles in various functions. Mutations in KH domains have been associated with multiple diseases, including fragile X syndrome. By characterizing the folding mechanism of a specific KH domain, it was found that transient intermediate accumulates and the domain has the propensity to form amyloid-like aggregates.