1B99

3'-FLUORO-URIDINE DIPHOSPHATE BINDING TO NUCLEOSIDE DIPHOSPHATE KINASE

1B99 の概要

• エントリーDOI: 10.2210/pdb1b99/pdb
• 分子名称: PROTEIN (NUCLEOSIDE DIPHOSPHATE KINASE), 2',3'-DIDEOXY-3'-FLUORO-URIDIDINE-5'-DIPHOSPHATE, PYROPHOSPHATE 2-, ... (4 entities in total)
• 機能のキーワード: phosphotransferase, antiviral agent, fluorouridine, transferase
• 由来する生物種: Dictyostelium discoideum
• 細胞内の位置: Cytoplasm: P22887
• タンパク質・核酸の鎖数: 6
• 化学式量合計: 102167.96

構造登録者

Janin, J.,Xu, Y. (登録日: 1999-02-22, 公開日: 1999-06-28, 最終更新日: 2023-08-09)

主引用文献

Gonin, P.,Xu, Y.,Milon, L.,Dabernat, S.,Morr, M.,Kumar, R.,Lacombe, M.L.,Janin, J.,Lascu, I.
Catalytic mechanism of nucleoside diphosphate kinase investigated using nucleotide analogues, viscosity effects, and X-ray crystallography.
Biochemistry, 38:7265-7272, 1999

PubMed Abstract: Nucleoside diphosphate (NDP) kinases display low specificity with respect to the base moiety of the nucleotides and to the 2'-position of the ribose, but the 3'-hydroxyl is found to be important for catalysis. We report in this paper the enzymatic analysis of a series of derivatives of thymidine diphosphate (TDP) where the 3'-OH group was removed or replaced by fluorine, azido, and amino groups. With Dictyostelium NDP kinase, kcat decreases 15-200-fold from 1100 s-1 with TDP, and (kcat/Km)NDP decreases from 12 x 10(6) to 10(3) to 5 x 10(4) M-1 s-1, depending on the substrate. The poorest substrates are 3'-deoxyTDP and 3'-azido-3'-deoxyTDP, while the best modified substrates are 2',3'-dehydro-3'-deoxyTDP and 3'-fluoro-3'-deoxyTDP. In a similar way, 3'-fluoro-2',3'-dideoxyUDP was found to be a better substrate than 2',3'-dideoxyUDP, but a much poorer substrate than 2'-deoxyUDP. (kcat/Km)NDP is sensitive to the viscosity of the solution with TDP as the substrate but not with the modified substrates. To understand the poor catalytic efficiency of the modified nucleotides at a structural level, we determined the crystal structure of Dictyostelium NDP kinase complexed to 3'-fluoro-2',3'-dideoxyUDP at 2.7 A resolution. Significant differences are noted as compared to the TDP complex. Substrate-assisted catalysis by the 3'-OH, which is effective in the NDP kinase reaction, cannot occur with the modified substrate. With TDP, the beta-phosphate, which is the leaving group when a gamma-phosphate is transferred to His122, hydrogen bonds to the 3'-hydroxyl group of the sugar; with 3'-fluoro-2',3'-dideoxyUDP, the beta-phosphate hydrogen bonds to Asn119 and moves away from the attacking Ndelta of the catalytic His122. Since all anti-AIDS nucleoside drugs are modified at the 3'-position, these results are relevant to the role of NDP kinase in their cellular metabolism.

PubMed: 10353838
DOI: 10.1021/bi982990v

実験手法

X-RAY DIFFRACTION (2.7 Å)