1OZF

The crystal structure of Klebsiella pneumoniae acetolactate synthase with enzyme-bound cofactors

1OZF の概要

• エントリーDOI: 10.2210/pdb1ozf/pdb
• 分子名称: Acetolactate synthase, catabolic, PHOSPHATE ION, MAGNESIUM ION, ... (6 entities in total)
• 機能のキーワード: acetolactate synthase, acetohydroxyacid synthase, thiamin diphosphate, lyase
• 由来する生物種: Klebsiella pneumoniae
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 124755.79

構造登録者

Pang, S.S.,Duggleby, R.G.,Schowen, R.L.,Guddat, L.W. (登録日: 2003-04-09, 公開日: 2003-11-11, 最終更新日: 2023-08-16)

主引用文献

Pang, S.S.,Duggleby, R.G.,Schowen, R.L.,Guddat, L.W.
The Crystal Structures of Klebsiella pneumoniae Acetolactate Synthase with Enzyme-bound Cofactor and with an Unusual Intermediate.
J.Biol.Chem., 279:2242-2253, 2004

PubMed Abstract: Acetohydroxyacid synthase (AHAS) and acetolactate synthase (ALS) are thiamine diphosphate (ThDP)-dependent enzymes that catalyze the decarboxylation of pyruvate to give a cofactor-bound hydroxyethyl group, which is transferred to a second molecule of pyruvate to give 2-acetolactate. AHAS is found in plants, fungi, and bacteria, is involved in the biosynthesis of the branched-chain amino acids, and contains non-catalytic FAD. ALS is found only in some bacteria, is a catabolic enzyme required for the butanediol fermentation, and does not contain FAD. Here we report the 2.3-A crystal structure of Klebsiella pneumoniae ALS. The overall structure is similar to AHAS except for a groove that accommodates FAD in AHAS, which is filled with amino acid side chains in ALS. The ThDP cofactor has an unusual conformation that is unprecedented among the 26 known three-dimensional structures of nine ThDP-dependent enzymes, including AHAS. This conformation suggests a novel mechanism for ALS. A second structure, at 2.0 A, is described in which the enzyme is trapped halfway through the catalytic cycle so that it contains the hydroxyethyl intermediate bound to ThDP. The cofactor has a tricyclic structure that has not been observed previously in any ThDP-dependent enzyme, although similar structures are well known for free thiamine. This structure is consistent with our proposed mechanism and probably results from an intramolecular proton transfer within a tricyclic carbanion that is the true reaction intermediate. Modeling of the second molecule of pyruvate into the active site of the enzyme with the bound intermediate is consistent with the stereochemistry and specificity of ALS.

PubMed: 14557277
DOI: 10.1074/jbc.M304038200

実験手法

X-RAY DIFFRACTION (2.3 Å)