1R49

Human topoisomerase I (Topo70) double mutant K532R/Y723F

1R49 の概要

• エントリーDOI: 10.2210/pdb1r49/pdb
• 関連するPDBエントリー: 1A36 1EJ9 1K4S 1K4T 1LPQ 1TMC
• 分子名称: 5'-D(*AP*AP*AP*AP*AP*GP*AP*CP*TP*TP*AP*GP*AP*AP*AP*AP*AP*TP*TP*TP*TP*T)-3', 5'-D(P*AP*AP*AP*AP*AP*TP*TP*TP*TP*TP*CP*TP*AP*AP*GP*TP*CP*TP*TP*TP*TP*T)-3', DNA topoisomerase I, ... (4 entities in total)
• 機能のキーワード: protein, dna, topoisomerase i, isomerase-dna complex, isomerase/dna
• 由来する生物種: Homo sapiens (human)
• 細胞内の位置: Nucleus, nucleolus: P11387
• タンパク質・核酸の鎖数: 3
• 化学式量合計: 83675.90

構造登録者

Interthal, H.,Quigley, P.M.,Hol, W.G.,Champoux, J.J. (登録日: 2003-10-03, 公開日: 2003-12-16, 最終更新日: 2024-02-14)

主引用文献

Interthal, H.,Quigley, P.M.,Hol, W.G.,Champoux, J.J.
The role of lysine 532 in the catalytic mechanism of human topoisomerase I.
J.Biol.Chem., 279:2984-2992, 2004

PubMed Abstract: Based on co-crystal structures of human topoisomerase I with bound DNA, Lys(532) makes a minor groove contact with the strongly preferred thymidine residue at the site of covalent attachment (-1 position). Replacement of Lys(532) with either arginine or alanine has essentially no effect on the sequence preference of the enzyme, indicating that this interaction is not required for the preference for a T at the -1 position. Although both the cleavage and religation activities of the K532R mutant enzyme are reduced, cleavage is reduced to a greater extent than religation. The reverse is true for the K532A mutant enzyme with religation so impaired that the nicked intermediate accumulates during plasmid relaxation assays. Consistent with the shift in the cleavage religation equilibrium toward cleavage for the K532A mutant enzyme, expression of the mutant enzyme in Saccharomyces cerevisiae is cytotoxic, and thus this mutant enzyme mimics the effects of the anticancer drug camptothecin. Cleavage assays with the mutant enzymes using an oligonucleotide containing a 5'-bridging phosphorothiolate indicate that Lys(532) functions as a general acid during cleavage to protonate the leaving 5'-oxygen. It is possible that the contact with the -1 base is important during catalysis to provide positional rigidity to the active site. The corresponding residues in the vaccinia virus topoisomerase and the tyrosine recombinases may have similar critical roles in catalysis.

PubMed: 14594810
DOI: 10.1074/jbc.M309959200

実験手法

X-RAY DIFFRACTION (3.13 Å)