With the recent advances in the field of protein structural biology it has become evident that understanding the functional repertoire of a protein not only necessitates the information of the stable three-dimensional structure but also the inherent dynamics associated with it. Metamorphic proteins are one such class of proteins which show explicit dynamics as they can exist in equilibrium with multiple native conformation without the assistance of any ligand or cofactors. Although the native structures of a few metamorphic proteins are known, the mechanism by which they undergo interconversion without misfolding remains largely unknown. A comprehensive knowledge of interconversion in metamorphic proteins can help the development of designed metamorphic proteins to target diseases.
Lymphotactin is a metamorphic chemokine protein which exists in a dynamic equilibrium between α+β monomer (Ltnαβ) and an all-β dimer (Ltnβ2). In this study we aim to detect and characterize higher energy on-pathway intermediates that will enable us to have a better understanding of fold switching process in lymphotactin and metamorphic proteins in general.
Using CEST and CPMG experiments we detected an intermediate which is in equilibrium with Ltnαβ with an exchange rate of ~500s-1 and population of 2%. Furthermore, our findings from a concentration dependent CEST experiment revealed that the intermediate is a monomer. We use multinuclear CEST and CPMG experiments to obtain the chemical shifts of the backbone of the intermediate, which are then used to calculate a structure of the intermediate using CS-Rosetta. The resulting structure of the monomeric intermediate consists of anti-parallel β-strands which closely resemble the structure of one protomer of Ltnβ2, thus making it a chimera intermediate having features from both Ltnαβ and Ltnβ2. The presence of this intermediate in the free energy landscape strongly suggests that this might be the on-pathway intermediate to interconversion between Ltnαβ and Ltnβ2.