5IFK

Purine nucleoside phosphorylase

5IFK の概要

• エントリーDOI: 10.2210/pdb5ifk/pdb
• 分子名称: Purine nucleoside phosphorylase, HYPOXANTHINE (3 entities in total)
• 機能のキーワード: purine nucleoside phosphorylase hypoxanthine np-i family kluyveromyces lactis, transferase
• 由来する生物種: Kluyveromyces lactis (strain ATCC 8585 / CBS 2359 / DSM 70799 / NBRC 1267 / NRRL Y-1140 / WM37) (Yeast)
• タンパク質・核酸の鎖数: 3
• 化学式量合計: 103119.15

構造登録者

Thakur, K.G.,Priyanka, A. (登録日: 2016-02-26, 公開日: 2017-03-01, 最終更新日: 2024-03-20)

主引用文献

Mahor, D.,Priyanka, A.,Prasad, G.S.,Thakur, K.G.
Functional and Structural Characterization of Purine Nucleoside Phosphorylase from Kluyveromyces lactis and Its Potential Applications in Reducing Purine Content in Food
PLoS ONE, 11:e0164279-e0164279, 2016

PubMed Abstract: Consumption of foods and beverages with high purine content increases the risk of hyperuricemia, which causes gout and can lead to cardiovascular, renal, and other metabolic disorders. As patients often find dietary restrictions challenging, enzymatically lowering purine content in popular foods and beverages offers a safe and attractive strategy to control hyperuricemia. Here, we report structurally and functionally characterized purine nucleoside phosphorylase (PNP) from Kluyveromyces lactis (KlacPNP), a key enzyme involved in the purine degradation pathway. We report a 1.97 Å resolution crystal structure of homotrimeric KlacPNP with an intrinsically bound hypoxanthine in the active site. KlacPNP belongs to the nucleoside phosphorylase-I (NP-I) family, and it specifically utilizes 6-oxopurine substrates in the following order: inosine > guanosine > xanthosine, but is inactive towards adenosine. To engineer enzymes with broad substrate specificity, we created two point variants, KlacPNPN256D and KlacPNPN256E, by replacing the catalytically active Asn256 with Asp and Glu, respectively, based on structural and comparative sequence analysis. KlacPNPN256D not only displayed broad substrate specificity by utilizing both 6-oxopurines and 6-aminopurines in the order adenosine > inosine > xanthosine > guanosine, but also displayed reversal of substrate specificity. In contrast, KlacPNPN256E was highly specific to inosine and could not utilize other tested substrates. Beer consumption is associated with increased risk of developing gout, owing to its high purine content. Here, we demonstrate that KlacPNP and KlacPNPN256D could be used to catalyze a key reaction involved in lowering beer purine content. Biochemical properties of these enzymes such as activity across a wide pH range, optimum activity at about 25°C, and stability for months at about 8°C, make them suitable candidates for food and beverage industries. Since KlacPNPN256D has broad substrate specificity, a combination of engineered KlacPNP and other enzymes involved in purine degradation could effectively lower the purine content in foods and beverages.

PubMed: 27768715
DOI: 10.1371/journal.pone.0164279

実験手法

X-RAY DIFFRACTION (1.967 Å)