Structural and Functional Characterization of Cancer-Associated D-Dopachrome Tautomerase Mutants

D-dopachrome tautomerase (D-DT), a member of the macrophage migration inhibitory factor (MIF) superfamily, is an immunomodulatory protein with activity in inflammation, autoimmune disorders, cardiovascular diseases, and cancer. The primary functionality of D-DT is associated with activation of the type II cell surface receptor, CD74. Upon activation, the D-DT-CD74 axis triggers a cascade of signaling events that promote cell survival and proliferation. In multiple types of cancer, the activation of D-DT-CD74 axis has been shown to have a tumor-promotion effect. The goal of this study is to examine the pathophysiological significance of thirteen D-DT mutations that were identified by next-generation sequencing (NGS) methods and reported in the Catalogue of Somatic Mutations in Cancer (COSMIC). These mutations, which are all located at the C-terminal region of D-DT, were identified in various solid tumors. The structural, functional, and biophysical properties of the D-DT variants will be interrogated with x-ray crystallography, circular dichroism (CD) spectroscopy, enzymatic assays, extracellular signal-regulated kinase (ERK) phosphorylation experiments, and molecular dynamics (MD) simulations. We anticipate that the data obtained by this study will have mechanistic value, enriching our understanding about the role of D-DT mutations in human cancers. Given the previously reported role of the C-terminal region in molecular recognition, we also expect that our findings will provide information into the structure–activity relationship (SAR) of D-DT promoting the currently ongoing drug discovery efforts.