Active Adenine Phosphoribosyltransferase (APRT)

Adenine Phosphoribosyltransferase (APRT) is an enzyme involved in the purine nucleotide salvage pathway. It functions as a catalyst in the reaction between adenine and phosphoribosyl pyrophosphate (PRPP) to form AMP. Besides, Vascular Cell Adhesion Molecule 1 (VCAM1) has been identified as an interactor of APRT, thus a binding ELISA assay was conducted to detect the interaction of recombinant human APRT and recombinant human VCAM1. Briefly, APRT were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100uL were then transferred to VCAM1-coated microtiter wells and incubated for 2h at 37℃. Wells were washed with PBST and incubated for 1h with anti-APRT pAb, then aspirated and washed 3 times. After incubation with HRP labelled secondary antibody, wells were aspirated and washed 3 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37℃. Finally, add 50µL stop solution to the wells and read at 450nm immediately. The binding activity of of APRT and VCAM1 was shown in Figure 1, and this effect was in a dose dependent manner.