1GTV

CRYSTAL STRUCTURE OF MYCOBACTERIUM TUBERCULOSIS THYMIDYLATE KINASE COMPLEXED WITH THYMIDINE-5'-DIPHOSPHATE (TDP)

1GTV の概要

• エントリーDOI: 10.2210/pdb1gtv/pdb
• 関連するPDBエントリー: 1G3U 1GSI
• 分子名称: THYMIDYLATE KINASE, MAGNESIUM ION, THYMIDINE-5'-DIPHOSPHATE, ... (7 entities in total)
• 機能のキーワード: transferase, transferase (atp:tmp phosphotransferase), kinase
• 由来する生物種: MYCOBACTERIUM TUBERCULOSIS
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 47028.70

構造登録者

Ursby, T.,Weik, M.,Fioravanti, E.,Delarue, M.,Goeldner, M.,Bourgeois, D. (登録日: 2002-01-21, 公開日: 2002-03-28, 最終更新日: 2024-05-08)

主引用文献

Ursby, T.,Weik, M.,Fioravanti, E.,Delarue, M.,Goeldner, M.,Bourgeois, D.
Cryophotolysis of Caged Compounds: A Technique for Trapping Intermediate States in Protein Crystals
Acta Crystallogr.,Sect.D, 58:607-, 2002

PubMed Abstract: Caged compounds in combination with protein crystallography represent a valuable tool in studies of enzyme reaction intermediates. To date, photochemical triggering of reactions has been performed close to room temperature. Synchronous reaction initiation has only been achieved with enzymes of relatively slow turnover (<0.1 s(-1)) and caged compounds of high quantum yield. Here X-ray crystallography and microspectrophotometry were used to provide evidence that (nitrophenyl)ethyl (NPE) ester bonds can be photolyzed by UV light at cryotemperatures. NPE-caged ATP in flash-cooled crystals of Mycobacterium tuberculosis thymidylate kinase was photolyzed successfully at 100-150 K as assessed by the structural observation of ATP-dependent enzymatic conversion of TMP to TDP after temporarily warming the crystals to room temperature. A new method is proposed in which cryo-photolysis combined with temperature-controlled protein crystallography can be used to trap reaction intermediates even in some of the fastest enzymes and/or when only compounds of low quantum yield are available. Raising the temperature after cryophotolysis may allow a transition barrier to be passed and an intermediate to accumulate in the crystal. A comparable method has only been used so far with proteins displaying endogenous photosensitivity. The approach described here opens the way to studying the reaction mechanisms of a much larger number of crystalline enzymes. Furthermore, it is shown that X-ray-induced radiolysis of caged compounds occurs if high-intensity synchrotron beamlines are used. This caveat should be taken into account when deriving data-collection protocols. It could also be used potentially as a way to trigger reactions.

PubMed: 11914484
DOI: 10.1107/S0907444902002135

実験手法

X-RAY DIFFRACTION (1.55 Å)