How pyridoxal 5′‐phosphate differentially regulates human cytosolic and mitochondrial serine hydroxymethyltransferase oligomeric state

Abstract

Adaptive metabolic reprogramming gives cancer cells a proliferative advantage. Tumour cells extensively use glycolysis to sustain anabolism and produce serine, which not only refuels the one-carbon units necessary for the synthesis of nucleotide precursors and for DNA methylation, but also affects the cellular redox homeostasis. Given its central role in serine metabolism, serine hydroxymethyltransferase (SHMT), a pyridoxal 5 0 -phosphate (PLP)-dependent enzyme, is an attractive target for tumour chemotherapy. In humans, the cytosolic isoform (SHMT1) and the mitochondrial isoform (SHMT2) have distinct cellular roles, but high sequence identity and comparable catalytic properties, which may complicate development of successful therapeutic strategies. Here, we investigated how binding of the cofactor PLP controls the oligomeric state of the human isoforms. The fact that eukaryotic SHMTs are tetrameric proteins while bacterial SHMTs function as dimers may suggest that the quaternary assembly in eukaryotes provides an advantage to fine-tune SHMT function and differentially regulate intertwined metabolic fluxes, and may provide a tool to address the specificity problem. We determined the crystal structure of SHMT2, and compared it to the apo-enzyme structure, showing that PLP binding triggers a disorder-to-order transition accompanied by a large rigid-body movement of the two cofactor-binding domains. Moreover, we demonstrated that SHMT1 exists in solution as a tetramer, both in the absence and presence of PLP, while SHMT2 undergoes a dimer-to-tetramer transition upon PLP binding. These findings indicate an unexpected structural difference between the two human SHMT isoforms, which opens new perspectives for understanding their differing behaviours, roles or regulation mechanisms in response to PLP availability in vivo.

SHMT2 Desuccinylation by SIRT5 Drives Cancer Cell Proliferation

“…In general, most of SHMT2 proteins form homodimer in cells without enzymatic activity (2). To activate its activity, SHMT2 goes through a transformation process to form tetramers after binding PLP at the K280 site.…”

“…As most of SHMT2 proteins form homodimers in cells without enzymatic activity, they must form tetramers to become active forms (2). Therefore, we further investigated whether K280E, which lowered the enzymatic activity of SHMT2 (Fig.…”

“…Mitochondrial serine hydroxymethyltransferase (SHMT2), a pyridoxal phosphate (PLP) binding protein, catalyzes the cleavage of serine to glycine accompanied with the production of 5, 10-methylenetetrahydrofolate (5, 10-CH 2 -THF), which is an essential intermediate for purine biosynthesis (1)(2)(3). SHMT2 is a direct c-myc target gene for cell survival during hypoxia (4,5), and its expression drives glioma cell survival in ischemia (6).…”

Oligomeric State and Thermal Stability of Apo- and Holo- Human Ornithine δ-Aminotransferase

“…The results reveal that apo and holo-hOAT exhibit similar tertiary structure, as shown by their near-UV CD spectra. Changes of the dichroic bands in the near-UV region during the apo to holo transition in Fold Type I aminotransferases were previously associated with changes in the position/orientation of aromatic residues at or in proximity of the active site [33]. In this family, PLP coordination typically involves a planar π base stacking interaction between the PLP pyridine ring and a tryptophan indolic ring, along with an H-bond interaction between a tyrosine residue belonging to the neighboring subunit and the phosphate group of PLP.…”

“…Thus, at present, it is not easy to understand how PLP binding can impact the tetrameric structure of the enzyme. In this regard, a conformational change has been associated to the PLP-induced tetramerization of mitochondrial human serine hydroxymethyltransferase [33]. The authors have suggested that PLP binding to the apoform shifts the equilibrium from an “open” to a “close” conformation of the dimers promoting the tetramerization.…”