Tài liệu Báo cáo khoa học: Kinetics of dextran-independent a-(1 fi 3)-glucan synthesis by

Kinetics of dextran-independent a-(1 fi 3)-glucan synthesis

by Streptococcus sobrinus glucosyltransferase I

Hideyuki Komatsu1

, Yoshie Abe1

, Kazuyuki Eguchi1

, Hideki Matsuno1

, Yu Matsuoka1

, Takayuki

Sadakane1

, Tetsuyoshi Inoue2

, Kazuhiro Fukui2 and Takao Kodama1

1 Department of Bioscience and Bioinformatics, Kyushu Institute of Technology, Iizuka, Japan

2 Department of Oral Microbiology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Science, Japan

Introduction

Water-insoluble glucan, which is mainly composed of

a-(1 fi 3)-glucan, enhances the colonization of cario￾genic bacteria and promotes the formation of dental

plaque on smooth tooth surfaces [1]. Two glucos￾yltransferases (GTFs) or glucansucrases (GTF-S and

GTF-I, EC 2.4.1.5) from mutans streptococci are

responsible for the production of this polysaccharide

[1,2]. GTF-S and GTF-I catalyze the synthesis of

water-soluble a-(1 fi 6)-glucan and water-insoluble

a-(1 fi 3)-glucan, respectively. They share highly

homologous amino acid sequences and comprise

the N-terminal glucansucrase domain (GSd) and the

C-terminal glucan-binding domain (GBd) [3,4]. The

GSd catalyzes glucosyl transfer from sucrose to low

molecular mass acceptors such as water, isomaltose,

and maltose [5]. On the other hand, the GBd binds to

glucans such as dextran [6–8].

Unlike GTF-S, pre-existing dextran increases GTF-I

activity [9,10]. Although truncation of the GBd of

GTF-I by genetic engineering results in decreased

Keywords

enzyme kinetics; glucansucrase;

glucosyltransferase; mutans streptococci;

nigerooligosaccharide

Correspondence

H. Komatsu, Department of Bioscience and

Bioinformatics, Kyushu Institute of

Technology, Kawazu 680-4, Iizuka 820-8502,

Japan

Fax: +81 948 29 7801

Tel: +81 948 29 7845

E-mail: [email protected]

(Received 3 September 2010, revised 18

November 2010, accepted 25 November

2010)

doi:10.1111/j.1742-4658.2010.07973.x

Glucosyltransferase (GTF)-I from cariogenic Streptococcus sobrinus elon￾gates the a-(1 fi 3)-linked glucose polymer branches on the primer dextran

bound to the C-terminal glucan-binding domain. We investigated the GTF￾I-catalyzed glucan synthesis reaction in the absence of the primer dextran.

The time course of saccharide production during dextran-independent glu￾can synthesis from sucrose was analyzed. Fructose and glucose were first

produced by the sucrose hydrolysis. Leucrose was subsequently produced,

followed by insoluble glucan [a-(1 fi 3)-linked glucose polymers] after a

lag phase. High levels of intermediate nigerooligosaccharide series accumu￾lation were characteristically not observed during the lag phase. The results

from the enzymatic activity of the acceptor reaction for the nigerooligo￾saccharide with a degree of polymerization of 2–6 and methyl a-D-gluco￾pyranoside as a glucose analog indicate that the activity increased with an

increase in the degree of polymerization. The production of insoluble glu￾can was numerically simulated using the fourth-order Runge–Kutta

method with the kinetic parameters estimated from the enzyme assay. The

simulated time course provided a profile similar to that of experimental

data. These results define the relationship between the kinetic properties

of GTF-I and the time course of saccharide production. These results are

discussed with respect to a mechanism that underlies efficient glucan

synthesis.

Abbreviations

DP, degree of polymerization; GBd, glucan-binding domain; GS, glucan-binding domain-deficient glucosyltransferase-I; GSd, glucansucrase

domain; GSGB, glucosyltransferase-I containing a full-length glucan-binding domain; GTF, glucosyltransferase.

FEBS Journal 278 (2011) 531–540 ª 2010 The Authors Journal compilation ª 2010 FEBS 531