Identification of a putative acetyltransferase gene, MMP0350, which affects proper assembly of both flagella and pili in the archaeon Methanococcus maripaludis

David J. VanDyke, John Wu, Sandy Y.M. Ng, Masaomi Kanbe, Bonnie Chaban, Shin Ichi Aizawa, Ken F. Jarrell

研究成果: 雜誌貢獻文章

37 引文 (Scopus)

摘要

Glycosylation is a posttranslational modification utilized in all three domains of life. Compared to eukaryotic and bacterial systems, knowledge of the archaeal processes involved in glycosylation is limited. Recently, Methanococcus voltae flagellin proteins were found to have an N-linked trisaccharide necessary for proper flagellum assembly. Current analysis by mass spectrometry of Methanococcus maripaludis flagellin proteins also indicated the attachment of an N-glycan containing acetylated sugars. To identify genes involved in sugar biosynthesis in M. maripaludis, a putative acetyltransferase was targeted for in-frame deletion. Deletion of this gene (MMP0350) resulted in a flagellin molecular mass shift to a size comparable to that expected for underglycosylated or completely nonglycoslyated flagellins, as determined by immunoblotting. Assembled flagellar filaments were not observed by electron microscopy. Interestingly, the deletion also resulted in defective pilus anchoring. Mutant cells with a deletion of MMP0350 had very few, if any, pili attached to the cell surface compared to a nonflagellated but piliated strain. However, pili were obtained from culture supernatants of this strain, indicating that the defect was not in pilus assembly but in stable attachment to the cell surface. Complementation of MMP0350 on a plasmid restored pilus attachment, but it was unable to restore flagellation, likely because the mutant ceased to make detectable flagellin. These findings represent the first report of a biosynthetic gene involved in flagellin glycosylation in archaea. Also, it is the first gene to be associated with pili, linking flagellum and pilus structure and assembly through posttranslational modifications.
原文英語
頁(從 - 到)5300-5307
頁數8
期刊Journal of Bacteriology
190
發行號15
DOIs
出版狀態已發佈 - 八月 1 2008
對外發佈Yes

指紋

Methanococcus
Flagellin
Acetyltransferases
Flagella
Archaea
Glycosylation
Genes
Post Translational Protein Processing
Trisaccharides
Gene Deletion
Immunoblotting
Polysaccharides
Mass Spectrometry
Electron Microscopy
Proteins
Plasmids

ASJC Scopus subject areas

  • Microbiology
  • Molecular Biology

引用此文

Identification of a putative acetyltransferase gene, MMP0350, which affects proper assembly of both flagella and pili in the archaeon Methanococcus maripaludis. / VanDyke, David J.; Wu, John; Ng, Sandy Y.M.; Kanbe, Masaomi; Chaban, Bonnie; Aizawa, Shin Ichi; Jarrell, Ken F.

於: Journal of Bacteriology, 卷 190, 編號 15, 01.08.2008, p. 5300-5307.

研究成果: 雜誌貢獻文章

VanDyke, David J. ; Wu, John ; Ng, Sandy Y.M. ; Kanbe, Masaomi ; Chaban, Bonnie ; Aizawa, Shin Ichi ; Jarrell, Ken F. / Identification of a putative acetyltransferase gene, MMP0350, which affects proper assembly of both flagella and pili in the archaeon Methanococcus maripaludis. 於: Journal of Bacteriology. 2008 ; 卷 190, 編號 15. 頁 5300-5307.
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abstract = "Glycosylation is a posttranslational modification utilized in all three domains of life. Compared to eukaryotic and bacterial systems, knowledge of the archaeal processes involved in glycosylation is limited. Recently, Methanococcus voltae flagellin proteins were found to have an N-linked trisaccharide necessary for proper flagellum assembly. Current analysis by mass spectrometry of Methanococcus maripaludis flagellin proteins also indicated the attachment of an N-glycan containing acetylated sugars. To identify genes involved in sugar biosynthesis in M. maripaludis, a putative acetyltransferase was targeted for in-frame deletion. Deletion of this gene (MMP0350) resulted in a flagellin molecular mass shift to a size comparable to that expected for underglycosylated or completely nonglycoslyated flagellins, as determined by immunoblotting. Assembled flagellar filaments were not observed by electron microscopy. Interestingly, the deletion also resulted in defective pilus anchoring. Mutant cells with a deletion of MMP0350 had very few, if any, pili attached to the cell surface compared to a nonflagellated but piliated strain. However, pili were obtained from culture supernatants of this strain, indicating that the defect was not in pilus assembly but in stable attachment to the cell surface. Complementation of MMP0350 on a plasmid restored pilus attachment, but it was unable to restore flagellation, likely because the mutant ceased to make detectable flagellin. These findings represent the first report of a biosynthetic gene involved in flagellin glycosylation in archaea. Also, it is the first gene to be associated with pili, linking flagellum and pilus structure and assembly through posttranslational modifications.",
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AU - Wu, John

AU - Ng, Sandy Y.M.

AU - Kanbe, Masaomi

AU - Chaban, Bonnie

AU - Aizawa, Shin Ichi

AU - Jarrell, Ken F.

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AB - Glycosylation is a posttranslational modification utilized in all three domains of life. Compared to eukaryotic and bacterial systems, knowledge of the archaeal processes involved in glycosylation is limited. Recently, Methanococcus voltae flagellin proteins were found to have an N-linked trisaccharide necessary for proper flagellum assembly. Current analysis by mass spectrometry of Methanococcus maripaludis flagellin proteins also indicated the attachment of an N-glycan containing acetylated sugars. To identify genes involved in sugar biosynthesis in M. maripaludis, a putative acetyltransferase was targeted for in-frame deletion. Deletion of this gene (MMP0350) resulted in a flagellin molecular mass shift to a size comparable to that expected for underglycosylated or completely nonglycoslyated flagellins, as determined by immunoblotting. Assembled flagellar filaments were not observed by electron microscopy. Interestingly, the deletion also resulted in defective pilus anchoring. Mutant cells with a deletion of MMP0350 had very few, if any, pili attached to the cell surface compared to a nonflagellated but piliated strain. However, pili were obtained from culture supernatants of this strain, indicating that the defect was not in pilus assembly but in stable attachment to the cell surface. Complementation of MMP0350 on a plasmid restored pilus attachment, but it was unable to restore flagellation, likely because the mutant ceased to make detectable flagellin. These findings represent the first report of a biosynthetic gene involved in flagellin glycosylation in archaea. Also, it is the first gene to be associated with pili, linking flagellum and pilus structure and assembly through posttranslational modifications.

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