One protein many functions: the non-canonical interactions of SHMT1. The structural and functional characterization of SHMT1 interactions with RNA and in the de novo thymidylate synthesis complex

Winner of the Competition “Prize for PhD Thesis 2023” arranged by Sapienza University Press. Several enzymes once characterized for specific metabolic roles are now recognized to have more than one function. Some metabolic enzymes, called moonlighting proteins, engage in non-canonical interactions with other proteins or nucleic acids. Interestingly, not all newly identified RNA-binding proteins regulate RNA function; sometimes, RNA itself dictates protein function through a mechanism known as riboregulation. For instance, the catalytic activity of human serine hydroxymethyltransferase (SHMT1) is modulated by RNA. Moreover, SHMT1 plays a key role in assembling the de novo thymidylate synthesis complex (dTMP-SC) in the nucleus alongside thymidylate synthase (TYMS) and dihydrofolate reductase (DHFR). This thesis aims to explore SHMT1’s non-canonical functions, focusing on the molecular mechanisms behind riboregulation. I will present the first 3D structure of a metabolic enzyme in complex with RNA, solved using cryo-electron microscopy, and characterize the dTMP-SC, highlighting its transient formation in vitro and in the cytosol. This manuscript explores how RNA regulates metabolic enzymes and transient interactions that influence function.