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Title: Structural and Functional Analysis of a New Subfamily of Glycosyltransferases Required for Glycosylation of Serine-rich Streptococcal Adhesins

Abstract

Serine-rich repeat glycoproteins (SRRPs) are a growing family of bacterial adhesins found in many streptococci and staphylococci; they play important roles in bacterial biofilm formation and pathogenesis. Glycosylation of this family of adhesins is essential for their biogenesis. A glucosyltransferase (Gtf3) catalyzes the second step of glycosylation of a SRRP (Fap1) from an oral streptococcus, Streptococcus parasanguinis. Although Gtf3 homologs are highly conserved in SRRP-containing streptococci, they share minimal homology with functionally known glycosyltransferases. We report here the 2.3 {angstrom} crystal structure of Gtf3. The structural analysis indicates that Gtf3 forms a tetramer and shares significant structural homology with glycosyltransferases from GT4, GT5, and GT20 subfamilies. Combining crystal structural analysis with site-directed mutagenesis and in vitro glycosyltransferase assays, we identified residues that are required for UDP- or UDP-glucose binding and for oligomerization of Gtf3 and determined their contribution to the enzymatic activity of Gtf3. Further in vivo studies revealed that the critical amino acid residues identified by the structural analysis are crucial for Fap1 glycosylation in S. parasanguinis in vivo. Moreover, Gtf3 homologs from other streptococci were able to rescue the gtf3 knock-out mutant of S. parasanguinis in vivo and catalyze the sugar transfer to the modified SRRP substrate inmore » vitro, demonstrating the importance and conservation of the Gtf3 homologs in glycosylation of SRRPs. As the Gtf3 homologs only exist in SRRP-containing streptococci, we conclude that the Gtf3 homologs represent a unique subfamily of glycosyltransferases.« less

Authors:
; ; ; ; ; ; ; ;  [1]
  1. (UAB)
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
FOREIGNNIH
OSTI Identifier:
1021194
Resource Type:
Journal Article
Journal Name:
J. Biol. Chem.
Additional Journal Information:
Journal Volume: 286; Journal Issue: (30) ; 07, 2011; Journal ID: ISSN 0021-9258
Country of Publication:
United States
Language:
ENGLISH
Subject:
59 BASIC BIOLOGICAL SCIENCES; 60 APPLIED LIFE SCIENCES; AMINO ACIDS; BACTERIA; BIOSYNTHESIS; CRYSTAL STRUCTURE; CRYSTALLOGRAPHY; ENZYMES; FUNCTIONAL ANALYSIS; GLYCOPROTEINS; IN VITRO; IN VIVO; MUTAGENESIS; MUTANTS; PATHOGENESIS; RESIDUES; SACCHAROSE; STREPTOCOCCUS; SUBSTRATES

Citation Formats

Zhu, Fan, Erlandsen, Heidi, Ding, Lei, Li, Jingzhi, Huang, Ying, Zhou, Meixian, Liang, Xiaobo, Ma, Jinbiao, and Wu, Hui. Structural and Functional Analysis of a New Subfamily of Glycosyltransferases Required for Glycosylation of Serine-rich Streptococcal Adhesins. United States: N. p., 2011. Web. doi:10.1074/jbc.M110.208629.
Zhu, Fan, Erlandsen, Heidi, Ding, Lei, Li, Jingzhi, Huang, Ying, Zhou, Meixian, Liang, Xiaobo, Ma, Jinbiao, & Wu, Hui. Structural and Functional Analysis of a New Subfamily of Glycosyltransferases Required for Glycosylation of Serine-rich Streptococcal Adhesins. United States. doi:10.1074/jbc.M110.208629.
Zhu, Fan, Erlandsen, Heidi, Ding, Lei, Li, Jingzhi, Huang, Ying, Zhou, Meixian, Liang, Xiaobo, Ma, Jinbiao, and Wu, Hui. Fri . "Structural and Functional Analysis of a New Subfamily of Glycosyltransferases Required for Glycosylation of Serine-rich Streptococcal Adhesins". United States. doi:10.1074/jbc.M110.208629.
@article{osti_1021194,
title = {Structural and Functional Analysis of a New Subfamily of Glycosyltransferases Required for Glycosylation of Serine-rich Streptococcal Adhesins},
author = {Zhu, Fan and Erlandsen, Heidi and Ding, Lei and Li, Jingzhi and Huang, Ying and Zhou, Meixian and Liang, Xiaobo and Ma, Jinbiao and Wu, Hui},
abstractNote = {Serine-rich repeat glycoproteins (SRRPs) are a growing family of bacterial adhesins found in many streptococci and staphylococci; they play important roles in bacterial biofilm formation and pathogenesis. Glycosylation of this family of adhesins is essential for their biogenesis. A glucosyltransferase (Gtf3) catalyzes the second step of glycosylation of a SRRP (Fap1) from an oral streptococcus, Streptococcus parasanguinis. Although Gtf3 homologs are highly conserved in SRRP-containing streptococci, they share minimal homology with functionally known glycosyltransferases. We report here the 2.3 {angstrom} crystal structure of Gtf3. The structural analysis indicates that Gtf3 forms a tetramer and shares significant structural homology with glycosyltransferases from GT4, GT5, and GT20 subfamilies. Combining crystal structural analysis with site-directed mutagenesis and in vitro glycosyltransferase assays, we identified residues that are required for UDP- or UDP-glucose binding and for oligomerization of Gtf3 and determined their contribution to the enzymatic activity of Gtf3. Further in vivo studies revealed that the critical amino acid residues identified by the structural analysis are crucial for Fap1 glycosylation in S. parasanguinis in vivo. Moreover, Gtf3 homologs from other streptococci were able to rescue the gtf3 knock-out mutant of S. parasanguinis in vivo and catalyze the sugar transfer to the modified SRRP substrate in vitro, demonstrating the importance and conservation of the Gtf3 homologs in glycosylation of SRRPs. As the Gtf3 homologs only exist in SRRP-containing streptococci, we conclude that the Gtf3 homologs represent a unique subfamily of glycosyltransferases.},
doi = {10.1074/jbc.M110.208629},
journal = {J. Biol. Chem.},
issn = {0021-9258},
number = (30) ; 07, 2011,
volume = 286,
place = {United States},
year = {2011},
month = {9}
}