Truffault V, Coles M, Diercks T, Abelmann K, Eberhardt S, Lüttgen H, Bacher A, Kessler H
Institut für Organische Chemie und Biochemie, Technische Universität München, Lichtenbergstrasse 4, 85747 Garching, Germany.
J Mol Biol. 2001 Jun 15;309(4):949-60. doi: 10.1006/jmbi.2001.4683.
The structure of the amino-terminal domain of Escherichia coli riboflavin synthase (RiSy) has been determined by NMR spectroscopy with riboflavin as a bound ligand. RiSy is functional as a 75 kDa homotrimer, each subunit of which consists of two domains which share very similar sequences and structures. The N-terminal domain (RiSy-N; 97 residues) forms a 20 kDa homodimer in solution which binds riboflavin with high affinity. The structure features a six-stranded antiparallel beta-barrel with a Greek-key fold, both ends of which are closed by an alpha-helix. One riboflavin molecule is bound per monomer in a site at one end of the barrel which is comprised of elements of both monomers. The structure and ligand binding are similar to that of the FAD binding domains of ferrodoxin reductase family proteins. The structure provides insights into the structure of the whole enzyme, the organisation of the functional trimer and the mechanism of riboflavin synthesis. C48 from the N-terminal domain is identified as the free cysteine implicated in a nucleophilic role in the synthesis mechanism, while H102 from the C-terminal domains is also likely to play a key role. Both are invariant in all known riboflavin synthase sequences.
通过以核黄素作为结合配体的核磁共振光谱法，已确定了大肠杆菌核黄素合酶（RiSy）氨基末端结构域的结构。RiSy作为一种75 kDa的同三聚体发挥功能，其每个亚基由两个结构域组成，这两个结构域具有非常相似的序列和结构。N末端结构域（RiSy-N；97个残基）在溶液中形成一个20 kDa的同二聚体，它以高亲和力结合核黄素。该结构的特征是具有希腊钥匙折叠的六链反平行β桶，其两端由一个α螺旋封闭。每个单体在桶一端的一个位点结合一个核黄素分子，该位点由两个单体的元件组成。其结构和配体结合与铁氧化还原蛋白还原酶家族蛋白的FAD结合结构域相似。该结构为了解整个酶的结构、功能性三聚体的组织以及核黄素合成机制提供了见解。来自N末端结构域的C48被确定为在合成机制中起亲核作用的游离半胱氨酸，而来自C末端结构域的H102也可能起关键作用。两者在所有已知的核黄素合酶序列中都是不变的。
