The effect of ATP on the folding and function of human uridine phosphorylase 1

• Main Authors: Yu-Ting Huang, 黃瑜婷
• Other Authors: Pei-Fen Liu
• Format: Others
• Language: zh-TW
• Published: 2017
• Online Access: http://ndltd.ncl.edu.tw/handle/71451682194300400214

碩士 === 國立中興大學 === 食品暨應用生物科技學系所 === 105 === Human Uridine phosphorylase 1 (hUP1) is an important enzyme in pyrimidine salvage pathway. This enzyme catalyzes the reversible phosphorylysis of uridine to uracil and ribose-1-phosphate. Meanwhile, the function of this enzyme is critical in activating 5-fluorouracil for chemotherapy. Interestingly, it was recently found that ATP apparently destabilizes E. coli uridine phosphorylase. However, whether ATP induces similar effect on human uridine phosphorylase 1 is still unknown. Here, we carefully investigated the effect of ATP on the folding and function of human uridine phosphorylase 1. Our results showed that ATP apparently decreases the stability of hUP1, and this destabilizing effect is not only relevant to the concentration of ATP but also hUP1. ANS-binding experiment suggests that ATP possibly accumulates the partially unfolded intermediate state of hUP1 for this apparent destabilization. Moreover, activity assay indicates that increased concentration of ATP simultaneously decreases the protein stability and inhibits the enzymatic activity. We also observed that ATP accelerates the unfolding rate of hUP1 with specific two phases. These results imply that ATP changes the protein folding and inhibits the enzyme activity of hUP1. This effect is possibly related to the efficacy of 5-FU in cancer cells that is well known with high-level of ATP for unconstrained proliferation.