Tài liệu Báo cáo khoa học: Structural basis for cyclodextrin recognition by Thermoactinomyces

Structural basis for cyclodextrin recognition by

Thermoactinomyces vulgaris cyclo⁄maltodextrin-binding

protein

Takashi Tonozuka1

, Akiko Sogawa1

, Mitsugu Yamada2

, Naoki Matsumoto1

, Hiromi Yoshida2,3,

Shigehiro Kamitori2,3, Kazuhiro Ichikawa1

, Masahiro Mizuno1,*, Atsushi Nishikawa1 and

Yoshiyuki Sakano1

1 Department of Applied Biological Science, Tokyo University of Agriculture and Technology, Japan

2 Graduate School of Medicine, Kagawa University, Japan

3 Information Technology Center, Kagawa University, Japan

Cyclodextrins (CDs) are cyclic a-1,4-glucans, and the

central cavity of CDs can host a large number of che￾micals by hydrophobic interaction [1]. A thermophilic

actinomycete, Thermoactinomyces vulgaris R-47, pro￾duces two CD-hydrolyzing enzymes, TVA I [2] and

TVA II [3]. We have determined the crystal structures

Keywords

crystal structure; cyclodextrin; sugar-binding

protein; sugar transporter; Thermoactinomyces

vulgaris

Correspondence

T. Tonozuka, Department of Applied

Biological Science, Tokyo University of

Agriculture and Technology, 3-5-8 Saiwai￾cho, Fuchu, Tokyo 183–8509, Japan

Fax: +81 42 3675705

Tel: +81 42 3675702

E-mail: [email protected]

*Present address

Department of Chemistry and Material

Engineering, Shinshu University, Nagano,

Japan

Database

The atomic coordinates and structural fac￾tors described in this paper have been

deposited in the Protein Data Bank (http://

www.rcsb.org/) with the accession code

2DFZ

(Received 11 December 2006, revised 13

February 2007, accepted 21 February 2007)

doi:10.1111/j.1742-4658.2007.05753.x

The crystal structure of a Thermoactinomyces vulgaris cyclo ⁄ maltodextrin￾binding protein (TvuCMBP) complexed with c-cyclodextrin has been deter￾mined. Like Escherichia coli maltodextrin-binding protein (EcoMBP) and

other bacterial sugar-binding proteins, TvuCMBP consists of two domains,

an N- and a C-domain, both of which are composed of a central b-sheet

surrounded by a-helices; the domains are joined by a hinge region contain￾ing three segments. c-Cyclodextrin is located at a cleft formed by the two

domains. A common functional conformational change has been reported

in this protein family, which involves switching from an open form

to a sugar-transporter bindable form, designated a closed form. The

TvuCMBP–c-cyclodextrin complex structurally resembles the closed form

of EcoMBP, indicating that TvuCMBP complexed with c-cyclodextrin

adopts the closed form. The fluorescence measurements also showed that

the affinities of TvuCMBP for cyclodextrins were almost equal to those for

maltooligosaccharides. Despite having similar folds, the sugar-binding site

of the N-domain part of TvuCMBP and other bacterial sugar-binding pro￾teins are strikingly different. In TvuCMBP, the side-chain of Leu59 pro￾trudes from the N-domain part into the sugar-binding cleft and orients

toward the central cavity of c-cyclodextrin, thus Leu59 appears to play the

key role in binding. The cleft of the sugar-binding site of TvuCMBP is also

wider than that of EcoMBP. These findings suggest that the sugar-binding

site of the N-domain part and the wide cleft are critical in determining the

specificity of TvuCMBP for c-cyclodextrin.

Abbreviations

CD, cyclodextrin; EcoMBP, Escherichia coli maltodextrin-binding protein; Mol, molecule; SeMet, selenomethionine; TliTMBP, Thermococcus

litoralis trehalose ⁄ maltose-binding protein; TvuCMBP, Thermoactinomyces vulgaris cyclo ⁄ maltodextrin-binding protein.

FEBS Journal 274 (2007) 2109–2120 ª 2007 The Authors Journal compilation ª 2007 FEBS 2109