TopoGEN provides purified Human Tyrosyl DNA Phosphodiesterase (Tdp1) purified to homogeneity to a single 68 kDa band on SDS-PAGE. Tdp1 is catalytically active and will remove a terminal 3’ phosphotyrosyl group from a specifically designed oligo target site (oligo-Tyr). Additionally, TopoGEN scientists have demonstrated that Tdp1 will release topo I covalently attached to genomic DNA isolated from cells treated with camptothecin using TopoGEN’s In Vivo Link Kit (TG1021). Tdp1 is thought to be important in repair of topo/DNA adducts that form following topo-active drug therapy; therefore, there is considerable interest in identifying drugs that target Tdp1. Such drugs may work well in combination with topo targeting strategies.
Figure 1. SDS-PAGE of TopoGEN Purified Tdp1. The gel was intentionally overloaded with 20ug of proten to reveal minor breakdown products.
One unit of Tdp1 can release the terminal Tyrosine from 20 ng of substrate (oligo-Tyr) in 30 min at 37° C.
Release of topo I from the covalent complex is shown below.
Biochemical assays for Tdp1 can be carried out using an oligo nucleotide with a 3’ terminal tyrosine residue. Release of tyrosine from a 5’ 32P end labeled oligo is easily measured using a denaturing polyacrylamide gel or DNA sequencing gels. TopoGEN Provides a kit for Tdp1 assay (TG1004) that has key reagents and controls/protocols for setting up a complete assay for Tdp1.
This product is shipped on wet ice and should be stored at – 20°C.