Investigations of the Active Site of the Enzyme
5,10-Methenyltetrahydrofolate Synthetase (MTHFS)
Folates are vitamins that are essential to human health. There are many forms of folate and each of these forms plays an important role in cellular metabolism. Folinic acid is a storage form of folate that is converted into more active forms of folate by an enzyme called 5,10-methenyltetrahydrofolate synthetase (MTHFS). These active forms of folate participate in essential cellular functions including synthesis of DNA, synthesis of protein, synthesis of ATP, and DNA repair. MTHFS is also important as its activity on folinic acid is an important step in two forms of chemotherapy.
Like most enzymes, MTHFS is a protein. Proteins are made up of subunits called amino acids. The goal of our project is to better understand the function of MTHFS by elucidating the bonding contacts its amino acids make with its substrates (folinic acid and ATP). To accomplish this, one amino acid in the natural form of MTHFS is exchanged for another using a collection of techniques from the fields of chemistry and biology. This creates a mutant protein that is then characterized for structural and functional changes through the use of spectroscopy and kinetic analysis. Structural or functional changes in the mutant enzyme give clues as to the role of the changed amino acid. This process is repeated for several selected amino acids to gain a better understanding of the enzyme as a whole. The collected information will improve the understanding of this important enzyme and the understanding of current medical therapies.