Inhibition of Dihydrofolate Reductase from W. Bancrofti
Presentation Type
Poster
Faculty Advisor
Nina Goodey
Access Type
Event
Start Date
26-4-2024 12:45 PM
End Date
26-4-2024 1:44 PM
Description
Lymphatic filariasis, also known as elephantiasis, is an infectious disease that takes place when filarial parasites are transmitted to humans through mosquitoes. These filarial parasites are worms called W. bancrofti (Wb) and they affect the lives of millions of individuals across the world. The symptoms vary from swollen limbs to high running fevers but most importantly lymphatic damage, which results in elephantiasis of the legs, arms, genitals and breasts. Dihydrofolate reductase is an established drug target in the treatment of cancer and bacterial infections. This enzyme plays a role in thymidylate synthesis and therefore its inhibition impacts DNA replication, weakening or killing the organism. Our research focuses on determining whether the Wb DHFR could be a possible drug target for Wb worms, which cause filariasis. DHFR is an enzyme that catalyzes the reduction of dihydrofolate (DHF) to tetrahydrofolate (THF). THF is necessary for DNA synthesis and methylation. We expressed Wb DHFR in E. coli cells in the presence of ampicillin and purified the protein through two types of affinity chromatography. The first column is a methotrexate-agarose column and the second column is a Ni-NTA resin. The expression vector we use for Wb DHFR results in a 6-His tag, which binds to the Ni-NTA resin and the enzyme binds to methotrexate, which is a competitive inhibitor. The purified protein is analyzed via SDS-PAGE electrophoresis and stored at -80 degrees Celsius to avoid denaturation. This protein will be used in inhibition assay to test existing compounds as potential inhibitors of Wb DHFR.
Inhibition of Dihydrofolate Reductase from W. Bancrofti
Lymphatic filariasis, also known as elephantiasis, is an infectious disease that takes place when filarial parasites are transmitted to humans through mosquitoes. These filarial parasites are worms called W. bancrofti (Wb) and they affect the lives of millions of individuals across the world. The symptoms vary from swollen limbs to high running fevers but most importantly lymphatic damage, which results in elephantiasis of the legs, arms, genitals and breasts. Dihydrofolate reductase is an established drug target in the treatment of cancer and bacterial infections. This enzyme plays a role in thymidylate synthesis and therefore its inhibition impacts DNA replication, weakening or killing the organism. Our research focuses on determining whether the Wb DHFR could be a possible drug target for Wb worms, which cause filariasis. DHFR is an enzyme that catalyzes the reduction of dihydrofolate (DHF) to tetrahydrofolate (THF). THF is necessary for DNA synthesis and methylation. We expressed Wb DHFR in E. coli cells in the presence of ampicillin and purified the protein through two types of affinity chromatography. The first column is a methotrexate-agarose column and the second column is a Ni-NTA resin. The expression vector we use for Wb DHFR results in a 6-His tag, which binds to the Ni-NTA resin and the enzyme binds to methotrexate, which is a competitive inhibitor. The purified protein is analyzed via SDS-PAGE electrophoresis and stored at -80 degrees Celsius to avoid denaturation. This protein will be used in inhibition assay to test existing compounds as potential inhibitors of Wb DHFR.