Evidence for the attachment of Hsp150/Pir2 to the cell wall of through disulfide bridges

Moukadiri, Ismaïl; Zueco, Jesús

doi:10.1016/s1567-1356(01)00025-3

Cited by 25 publications

(32 citation statements)

References 13 publications

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“…Hsp150 and Cis3 were also found among the proteins released upon treatment of the cell wall with ␤-mercaptoethanol or dithiothreitol, showing that they are linked to other CWPs through disulfide bonds (Fig. 1A), probably involving the cysteine-rich C terminus (39,43,189).…”

Section: Mannoproteinsmentioning

confidence: 99%

Cell Wall Assembly in Saccharomyces cerevisiae

Lesage

Bussey

2006

Microbiol Mol Biol Rev

720

749

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SUMMARY An extracellular matrix composed of a layered meshwork of β-glucans, chitin, and mannoproteins encapsulates cells of the yeast Saccharomyces cerevisiae. This organelle determines cellular morphology and plays a critical role in maintaining cell integrity during cell growth and division, under stress conditions, upon cell fusion in mating, and in the durable ascospore cell wall. Here we assess recent progress in understanding the molecular biology and biochemistry of cell wall synthesis and its remodeling in S. cerevisiae. We then review the regulatory dynamics of cell wall assembly, an area where functional genomics offers new insights into the integration of cell wall growth and morphogenesis with a polarized secretory system that is under cell cycle and cell type program controls.

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Section: Mannoproteinsmentioning

confidence: 99%

Cell Wall Assembly in Saccharomyces cerevisiae

Lesage

Bussey

2006

Microbiol Mol Biol Rev

720

749

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“…Because Pir proteins can be liberated from the cell wall by mild alkaline extraction and because they are highly O glycosylated, it is thought that they are retained through an O-glycosidic linkage to ␤-1,3-glucan (9,19,29). It has also been reported that Pir proteins are bound to the cell wall by disulfide bridges because they can be released by a reducing agent such as ␤-mercaptoethanol or dithiothreitol (15,28,31). Furthermore, Castillo et al reported that the repetitive sequence of Pir4p is necessary for its binding to the cell wall (5).…”

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confidence: 99%

Comparison of Cell Wall Localization among Pir Family Proteins and Functional Dissection of the Region Required for Cell Wall Binding and Bud Scar Recruitment of Pir1p

et al. 2005

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We examined the localization of the Pir protein family (Pir1 to Pir4), which is covalently linked to the cell wall in an unknown manner. In contrast to the other Pir proteins, a fusion of Pir1p and monomeric red fluorescent protein distributed in clusters in pir1⌬ cells throughout the period of cultivation, indicating that Pir1p is localized in bud scars. Further microscopic analysis revealed that Pir1p is expressed inside the chitin rings of the bud scars. Stepwise deletion of the eight units of the repetitive sequence of Pir1p revealed that one unit is enough for the protein to bind bud scars and that the extent of binding of Pir1p to the cell wall depends on the number of these repetitive units. The localization of a chimeric Pir1p in which the repetitive sequence of Pir1p was replaced with that of Pir4p revealed the functional role of the different protein regions, specifically, that the repetitive sequence is required for binding to the cell wall and that the C-terminal sequence is needed for recruitment to bud scars. This is the first report that bud scars contain proteins like Pir1p as internal components.

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“…The PIR gene family is known to have two binding regions for the cell wall. One is the repetitive sequence, which is necessary for binding with ␤1,3-glucan in an alkaline-sensitive manner; the other is the cysteine residue-rich C-terminal region, which is necessary for binding with an unidentified component of the cell wall through the disulfide bond (12,34,51).…”

Section: Resultsmentioning

confidence: 99%

Construction of a Library of Human Glycosyltransferases Immobilized in the Cell Wall ofSaccharomyces cerevisiae

Shimma¹,

Saito²,

Oosawa³

et al. 2006

Appl Environ Microbiol

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Fifty-one human glycosyltransferases were expressed in Saccharomyces cerevisiae as immobilized enzymes and were assayed for enzymatic activities. The stem and catalytic regions of sialyl-, fucosyl-, galactosyl-, N-acetylgalactosaminyl-, and N-acetylglucosaminyltransferases were fused with yeast cell wall Pir proteins, which anchor glycosyltransferases at the yeast cell wall glucan. More than 75% of expressed recombinant glycosyltransferases retained their enzymatic activities in the yeast cell wall fraction and will be used as a human glycosyltransferase library. In increasing the enzymatic activities of immobilized glycosyltransferases, several approaches were found to be effective. Additional expression of yeast protein disulfide isomerase increased the expression levels and activities of polypeptide N-acetylgalactosaminyltransferases and other glycosyltransferases. PIR3 and/or PIR4 was more effective than PIR1 as a cell wall anchor when the Pirglycosyltransferase fusions were expressed under the control of the constitutive glyceraldehyde-3-phosphate dehydrogenase promoter. Oligosaccharides such as Lewis x, Lewis y, and H antigen were successfully synthesized using this immobilized glycosyltransferase library, indicating that the Pir-fused glycosyltransferases are useful for the production of various human oligosaccharides.Most oligosaccharides exist as glycoproteins and glycolipids, many of which are localized at the cell surface and are involved in biologically important processes such as cellular adhesion, molecular recognition, and signal transduction (58). The oligosaccharides are extremely rich in structural variations, and isomers exhibit different branching structures, anomeric configurations, and linkage positions; these vary widely with species, tissue, and degree of cellular differentiation. To clarify the structure-function relationship of oligosaccharides, the synthesis and remodeling of oligosaccharides are required. In order to synthesize complex oligosaccharides on proteins and lipids or in living cells, enzymatic methods are more suitable than chemical methods, since degradation or denaturation of active proteins, lipids, and living cells can be avoided. Human-derived enzymes are particularly ideal for the synthesis of mammalian oligosaccharides, such as sialylated and branched glycans, because these reactions are difficult to reproduce with either chemical methods or bacterial enzymes (19).At present, nearly 200 human glycosyltransferase genes are known and are available for use. These are indispensable for enzymatic synthesis of human-type oligosaccharides (41). However, it is important to make these resources suitable for application as enzyme sources. Immobilized enzymes enable the recovery of valuable enzymes for repeated usage and have several advantages for industrial applications; they are usually more stable than free enzymes and can be applied via automated bioreactor-packed columns.

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Evidence for the attachment of Hsp150/Pir2 to the cell wall of through disulfide bridges

Cited by 25 publications

References 13 publications

Cell Wall Assembly in Saccharomyces cerevisiae

Cell Wall Assembly in Saccharomyces cerevisiae

Comparison of Cell Wall Localization among Pir Family Proteins and Functional Dissection of the Region Required for Cell Wall Binding and Bud Scar Recruitment of Pir1p

Construction of a Library of Human Glycosyltransferases Immobilized in the Cell Wall ofSaccharomyces cerevisiae

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