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Infection and Immunity, December 2008, p. 5624-5631, Vol. 76, No. 12
0019-9567/08/$08.00+0     doi:10.1128/IAI.00534-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

A Conserved C-Terminal 13-Amino-Acid Motif of Gap1 Is Required for Gap1 Function and Necessary for the Biogenesis of a Serine-Rich Glycoprotein of Streptococcus parasanguinis

Meixian Zhou,¹ Zhixiang Peng,¹ Paula Fives-Taylor,² and Hui Wu^1*

Departments of Pediatric Dentistry and Microbiology, University of Alabama at Birmingham Schools of Medicine and Dentistry, Birmingham, Alabama 35244,¹ Department of Microbiology and Molecular Genetics, College of Medicine and College of Life Science and Agriculture, Burlington, Vermont 05405²

Received 30 April 2008/ Returned for modification 17 May 2008/ Accepted 29 September 2008

Adhesion of Streptococcus parasanguinis to saliva-coated hydroxyapatite (SHA), an in vitro tooth model, is mediated by long peritrichous fimbriae. Fap1, a fimbria-associated serine-rich glycoprotein, is required for fimbrial assembly. Biogenesis of Fap1 is controlled by an 11-gene cluster that contains gly, nss, galT1 and -2, secY2, gap1 to -3, secA2, and gtf1 and -2. We had previously isolated a collection of nine nonadherent mutants using random chemical mutagenesis approaches. These mutants fail to adhere to the in vitro tooth model and to form fimbriae. In this report, we further characterized these randomly selected nonadherent mutants and classified them into three distinct groups. Two groups of genes were previously implicated in Fap1 biogenesis. One group has a mutation in a glycosyltransferase gene, gtf1, that is essential for the first step of Fap1 glycosylation, whereas the other group has defects in the fap1 structural gene. The third group mutant produces an incompletely glycosylated Fap1 and exhibits a mutant phenotype similar to that of a glycosylation-associated protein 1 (Gap1) mutant. Analysis of this new mutant revealed that a conserved C-terminal 13-amino-acid motif was missing in Gap1. Site-directed mutagenesis of a highly conserved amino acid tryptophan within this motif recapitulated the deletion phenotype, demonstrating the importance of the Gap1 C-terminal motif for Fap1 biogenesis. Furthermore, the C-terminal mutation does not affect Gap1-Gap3 protein-protein interaction, which has been shown to mediate Fap1 glycosylation, suggesting the C-terminal motif has a distinct function related to Fap1 biogenesis.

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* Corresponding author. Mailing address: Department of Pediatric Dentistry, University of Alabama at Birmingham School of Dentistry, Birmingham, AL 35244. Phone: (205) 996-2392. Fax: (205) 975-6251. E-mail: hwu{at}uab.edu

Published ahead of print on 13 October 2008.

Editor: A. Camilli

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Infection and Immunity, December 2008, p. 5624-5631, Vol. 76, No. 12
0019-9567/08/$08.00+0     doi:10.1128/IAI.00534-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

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