8RH3

Nucleoside 2'deoxyribosyltransferase from Chroococcidiopsis thermalis PCC 7203 WT bound to Gemcitabine

これはPDB形式変換不可エントリーです。

8RH3 の概要

• エントリーDOI: 10.2210/pdb8rh3/pdb
• 分子名称: Nucleoside 2-deoxyribosyltransferase, (2~{R},3~{R})-4,4-bis(fluoranyl)-2-(hydroxymethyl)oxolan-3-ol (3 entities in total)
• 機能のキーワード: ribosylated, holoprotein, transferase, gemcitabine
• 由来する生物種: Chroococcidiopsis thermalis PCC 7203
• タンパク質・核酸の鎖数: 4
• 化学式量合計: 72126.03

構造登録者

Tang, P.,Harding, C.J.,Czekster, C.M. (登録日: 2023-12-14, 公開日: 2024-02-21, 最終更新日: 2024-11-13)

主引用文献

Tang, P.,Harding, C.J.,Dickson, A.L.,da Silva, R.G.,Harrison, D.J.,Czekster, C.M.
Snapshots of the Reaction Coordinate of a Thermophilic 2'-Deoxyribonucleoside/ribonucleoside Transferase.
Acs Catalysis, 14:3090-3102, 2024

PubMed Abstract: Nucleosides are ubiquitous to life and are required for the synthesis of DNA, RNA, and other molecules crucial for cell survival. Despite the notoriously difficult organic synthesis of nucleosides, 2'-deoxynucleoside analogues can interfere with natural DNA replication and repair and are successfully employed as anticancer, antiviral, and antimicrobial compounds. Nucleoside 2'-deoxyribosyltransferase (dNDT) enzymes catalyze transglycosylation via a covalent 2'-deoxyribosylated enzyme intermediate with retention of configuration, having applications in the biocatalytic synthesis of 2'-deoxynucleoside analogues in a single step. Here, we characterize the structure and function of a thermophilic dNDT, the protein from (NDT). We combined enzyme kinetics with structural and biophysical studies to dissect mechanistic features in the reaction coordinate, leading to product formation. Bell-shaped pH-rate profiles demonstrate activity in a broad pH range of 5.5-9.5, with two very distinct p values. A pronounced viscosity effect on the turnover rate indicates a diffusional step, likely product (nucleobase1) release, to be rate-limiting. Temperature studies revealed an extremely curved profile, suggesting a large negative activation heat capacity. We trapped a 2'-fluoro-2'-deoxyarabinosyl-enzyme intermediate by mass spectrometry and determined high-resolution structures of the protein in its unliganded, substrate-bound, ribosylated, 2'-difluoro-2'-deoxyribosylated, and in complex with probable transition-state analogues. We reveal key features underlying (2'-deoxy)ribonucleoside selection, as NDT can also use ribonucleosides as substrates, albeit with a lower efficiency. Ribonucleosides are the building blocks of RNA and other key intracellular metabolites participating in energy and metabolism, expanding the scope of use of NDT in biocatalysis.

PubMed: 38449528
DOI: 10.1021/acscatal.3c06260

実験手法

X-RAY DIFFRACTION (1.96 Å)