Global Fold Switching of the RafH Protein: Diverse Structures with a Conserved Pathway.

ABSTRACT

It is generally believed that a protein's sequence uniquely determines its structure, the basis for a protein to perform biological functions. However, as a representative metamorphic protein, RfaH can be encoded by a single amino acid sequence into two distinct native state structures. Its C-terminal domain (CTD) either takes an all-α-helical configuration to pack tightly with its N-terminal domain (NTD), or the CTD disassociates from the NTD, transforms into an all-ß-barrel fold, and further attaches to the ribosome, leaving the NTD exposed to bind RNA polymerases. Therefore, the RfaH protein couples transcription and translation processes. Although previous studies have provided a preliminary understanding of its function, the full course of the conformational change of RfaH-CTD at the atomic level is elusive. We used teDA2, a feature space-based enhanced sampling protocol, to explore the transformation of RfaH-CTD. We found that it undergoes a large-scale structural rearrangement, with characteristic spectra as the fingerprint, and a global unfolding transition with a tighter and energetically moderate molten globule-like nucleus formed in between. The formation of this nucleus limits the possible intermediate conformations, facilitates the formation of secondary and tertiary structures, and thus ensures the efficiency of transformation. The key features along the transition path disclosed from this work are likely associated with the evolution of RfaH, such that encoding a single sequence into multiple folds with distinct biological functions is energetically unhindered.