Engineering Sulfatases for Biomedical Applications
Heparin, which is obtained from pig tissue, has been used as an anti-coagulant for a century. However, a contamination scare in 2008, which caused fatalities worldwide, has amplified the need for homogenous synthetic heparins. Given the difficulties often associated with chemical synthesis of large polysaccharides, enzymatic approaches are much more tractable. Ultra low molecular weight heparins (ULMWHs) have been demonstrated to work effectively as anti-coagulants. However, ULMWHs have one drawback as they can't be neutralized by protamine sulfate, meaning there is no antidote if a patient is given an overdose of anti-coagulant. The aim of this project is to develop a robust assay to measure G6S activity, using novel synthetic substrates that will be synthesized as part of this project. This, in addition to the structural information already obtained, will be used as the basis for engineering G6S, using rational site directed mutagenesis, in order to obtain an enzyme that will work at higher pH. Such an engineered enzyme could then be explored for use therapeutically as an antidote to anti-coagulant overdose, and scale-up of protein expression for an industrial application will be addressed.
3/6 complete
Imported from UKRI Gateway to Research | Grant 13D7C04A-A7F2-4916-839D-003985589FB7.
Open sourceOverview
Grant agreement 13D7C04A-A7F2-4916-839D-003985589FB7
Open sourceHeparin, which is obtained from pig tissue, has been used as an anti-coagulant for a century. However, a contamination scare in 2008, which caused fatalities worldwide, has amplified the need for homogenous synthetic heparins. Given the difficulties often associated with chemical synthesis of large polysaccharides, enzymatic approaches are much more tractable. Ultra low molecular weight heparins (ULMWHs) have been demonstrated to work effectively as anti-coagulants. However, ULMWHs have one drawback as they can't be neutralized by protamine sulfate, meaning there is no antidote if a patient is given an overdose of anti-coagulant. The aim of this project is to develop a robust assay to measure G6S activity, using novel synthetic substrates that will be synthesized as part of this project. This, in addition to the structural information already obtained, will be used as the basis for engineering G6S, using rational site directed mutagenesis, in order to obtain an enzyme that will work at higher pH. Such an engineered enzyme could then be explored for use therapeutically as an antidote to anti-coagulant overdose, and scale-up of protein expression for an industrial application will be addressed.