Abstract
Glucosyltransferase-I (GTF-I: 175 kDa) of a cariogenic bacterium, Streptococcus sobrinus 6715, mediates the conversion of water-soluble dextran (α-1,6-glucan) into a water-insoluble form by making numerous α-1,3-glucan branches along the dextran chains with sucrose as the glucosyl donor. The structures and catalytic properties were compared for two GTF-I fragments, GTF-I' (138 kDa) and GS (110 kDa). Both lack the N-terminal 84 residues of GTF-I. While GTF-I' still contains four of the six C-terminal repeats characteristic of streptococcal glucosyltransferases, GS lacks all of them. Electron microscopy of negatively stained samples indicated a double-domain structure for GTF-I', consisting of a spherical head with a smaller spherical tail, which was occasionally seen as a long extension. GS was seen just as the head portion of GTF-I'. In the absence of dextran, both fragments simply hydrolyzed sucrose with similar K(m) and k(cat) values at low concentrations (<5 mM). At higher sucrose concentrations (> 10 mM), however, GTF-I' exhibited glucosyl transfer activity to form insoluble α-1,3-glucans. So did GS, but less efficiently. Dextran increased the rate and efficiency of the glucosyl transfer by GTF-I'. On removal of the C-terminal repeats of GTF-I' by mild trypsin treatment, this dextran-stimulated transfer was completely lost and the dextran-independent transfer became less efficient. These results indicate that the N-terminal two-thirds of the GTF-I sequence are organized as a structurally and functionally independent domain to catalyze not only sucrose hydrolysis but also glucosyl transfer to form α-1,3-glucan chains, although not efficiently; the C-terminal repeat increases the efficiency of the intrinsic glucosyl transfer by the N-terminal domain as well as rendering the whole molecule primer-dependent for far more efficient insoluble glucan synthesis.
| Original language | English |
|---|---|
| Pages (from-to) | 287-295 |
| Number of pages | 9 |
| Journal | Journal of Biochemistry |
| Volume | 126 |
| Issue number | 2 |
| Publication status | Published - 1999 |
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Keywords
- α-1,3-glucan
- C-terminal repeats
- Dextran
- Domain structure
- Glucosyltransferase
ASJC Scopus subject areas
- Biochemistry
Cite this
Structure and enzymatic properties of genetically truncated forms of the water-insoluble glucan-synthesizing glucosyltransferase from Streptococcus sobrinus. / Konishi, Norifumi; Torii, Yasuhiro; Yamamoto, Tatsuo; Miyagi, Atsushi; Ohta, Hiroyuki; Fukui, Kazuhiro; Hanamoto, Satoshi; Matsuno, Hideki; Komatsu, Hideyuki; Kodama, Takao; Katayama, Eisaku.
In: Journal of Biochemistry, Vol. 126, No. 2, 1999, p. 287-295.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Structure and enzymatic properties of genetically truncated forms of the water-insoluble glucan-synthesizing glucosyltransferase from Streptococcus sobrinus
AU - Konishi, Norifumi
AU - Torii, Yasuhiro
AU - Yamamoto, Tatsuo
AU - Miyagi, Atsushi
AU - Ohta, Hiroyuki
AU - Fukui, Kazuhiro
AU - Hanamoto, Satoshi
AU - Matsuno, Hideki
AU - Komatsu, Hideyuki
AU - Kodama, Takao
AU - Katayama, Eisaku
PY - 1999
Y1 - 1999
N2 - Glucosyltransferase-I (GTF-I: 175 kDa) of a cariogenic bacterium, Streptococcus sobrinus 6715, mediates the conversion of water-soluble dextran (α-1,6-glucan) into a water-insoluble form by making numerous α-1,3-glucan branches along the dextran chains with sucrose as the glucosyl donor. The structures and catalytic properties were compared for two GTF-I fragments, GTF-I' (138 kDa) and GS (110 kDa). Both lack the N-terminal 84 residues of GTF-I. While GTF-I' still contains four of the six C-terminal repeats characteristic of streptococcal glucosyltransferases, GS lacks all of them. Electron microscopy of negatively stained samples indicated a double-domain structure for GTF-I', consisting of a spherical head with a smaller spherical tail, which was occasionally seen as a long extension. GS was seen just as the head portion of GTF-I'. In the absence of dextran, both fragments simply hydrolyzed sucrose with similar K(m) and k(cat) values at low concentrations (<5 mM). At higher sucrose concentrations (> 10 mM), however, GTF-I' exhibited glucosyl transfer activity to form insoluble α-1,3-glucans. So did GS, but less efficiently. Dextran increased the rate and efficiency of the glucosyl transfer by GTF-I'. On removal of the C-terminal repeats of GTF-I' by mild trypsin treatment, this dextran-stimulated transfer was completely lost and the dextran-independent transfer became less efficient. These results indicate that the N-terminal two-thirds of the GTF-I sequence are organized as a structurally and functionally independent domain to catalyze not only sucrose hydrolysis but also glucosyl transfer to form α-1,3-glucan chains, although not efficiently; the C-terminal repeat increases the efficiency of the intrinsic glucosyl transfer by the N-terminal domain as well as rendering the whole molecule primer-dependent for far more efficient insoluble glucan synthesis.
AB - Glucosyltransferase-I (GTF-I: 175 kDa) of a cariogenic bacterium, Streptococcus sobrinus 6715, mediates the conversion of water-soluble dextran (α-1,6-glucan) into a water-insoluble form by making numerous α-1,3-glucan branches along the dextran chains with sucrose as the glucosyl donor. The structures and catalytic properties were compared for two GTF-I fragments, GTF-I' (138 kDa) and GS (110 kDa). Both lack the N-terminal 84 residues of GTF-I. While GTF-I' still contains four of the six C-terminal repeats characteristic of streptococcal glucosyltransferases, GS lacks all of them. Electron microscopy of negatively stained samples indicated a double-domain structure for GTF-I', consisting of a spherical head with a smaller spherical tail, which was occasionally seen as a long extension. GS was seen just as the head portion of GTF-I'. In the absence of dextran, both fragments simply hydrolyzed sucrose with similar K(m) and k(cat) values at low concentrations (<5 mM). At higher sucrose concentrations (> 10 mM), however, GTF-I' exhibited glucosyl transfer activity to form insoluble α-1,3-glucans. So did GS, but less efficiently. Dextran increased the rate and efficiency of the glucosyl transfer by GTF-I'. On removal of the C-terminal repeats of GTF-I' by mild trypsin treatment, this dextran-stimulated transfer was completely lost and the dextran-independent transfer became less efficient. These results indicate that the N-terminal two-thirds of the GTF-I sequence are organized as a structurally and functionally independent domain to catalyze not only sucrose hydrolysis but also glucosyl transfer to form α-1,3-glucan chains, although not efficiently; the C-terminal repeat increases the efficiency of the intrinsic glucosyl transfer by the N-terminal domain as well as rendering the whole molecule primer-dependent for far more efficient insoluble glucan synthesis.
KW - α-1,3-glucan
KW - C-terminal repeats
KW - Dextran
KW - Domain structure
KW - Glucosyltransferase
UR - http://www.scopus.com/inward/record.url?scp=0032818249&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0032818249&partnerID=8YFLogxK
M3 - Article
VL - 126
SP - 287
EP - 295
JO - Journal of Biochemistry
JF - Journal of Biochemistry
SN - 0021-924X
IS - 2
ER -