Measurement of Hydrolytic Activity of DFsc Proteins Using 4-Methylumbelliferyl Phosphate and Bis(4-nitrophenyl) Phosphate
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Many proteins that are found in nature contain metals that help perform catalytic functions. Metallohydrolases are one such class of enzymes that cleave chemical bonds using water and are responsible for important biological functions such as oxygen reduction and photosynthesis. One method to better understand the function of these proteins is to use a synthetic model system. Due Ferri single chain proteins (DFsc), a family of synthetic model proteins first used to study diiron enzymes has an active site similar to the metallohydrolases found in nature. To see if DFsc can function as a metallohydrolase, the colorimetric probe bis-(4-nitrophenyl) phosphate (BNPP) was used to monitor hydrolytic catalysis in the presence of manganese (II) and zinc (II) ions. Both Mn(II)- and Zn(II)-bound DFsc proteins exhibited catalytic activity at pH 7.4 and 7.8. The effect of active site mutants was also explored by comparing rates of G4DFsc to DFsc, with G4DFsc showing faster hydrolytic cleavage rates. Understanding how the DFsc class of proteins perform hydrolysis reactions can help explain the reactivity in natural metallohydrolases, as well as produce new metallohydrolases that can be used in pharmaceutical and biotechnological areas.
Pantel, Joseph, "Measurement of Hydrolytic Activity of DFsc Proteins Using 4-Methylumbelliferyl Phosphate and Bis(4-nitrophenyl) Phosphate" (2021). Chemistry Honors Papers. 67.