Serine hydroxymethyltransferase is an enzyme that catalyzes the reversible interconversion of the amino acids serine and glycine using the one-carbon carrier tetrahydrofolate and pyridoxal phosphate (PLP), the active form of vitamin B6.
The enzyme purified from E. coli is a 95 kDa homodimer. The enzyme mechanism has been well-characterized, and the crystal structure determined. One area of current research focuses on the folding mechanism and how the enzyme obtains the PLP cofactor.
SHMT Folding Studies
Denaturation of SHMT with urea, followed by dilution, allows complete renaturation of the enzyme and restoration of activity. Renaturation experiments using using single tryptophan mutants and fluorescence spectrometry show that the PLP cofactor is not added until after dimer formation, even though the active sites are buried at the interface between the two subunits. Therefore, some unknown folding mechanism must put the enzyme in a different conformation that gives the active site access to the PLP, either from the solvent or via channeling from another enzyme.
Several SHMT glycine residues are highly conserved in SHMT primary sequences from diverse species. Analysis of three-dimensional structures from those same enzymes, done in this lab, confirmed that the glycine residues occupy homologous positions. As is commonly the case, those glycine residues are located at turns in the enyzme tertiary structures. The greater flexibility of glycine residue often allows them to function as hinge points.
In collaboration with other laboratories, a series of site-directed mutants of SHMT have been prepared with the key glycine residues substituted for bulkier valines. GC students in this lab are characterizing the kinetic and folding properties of those mutant SHMTs.