| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
Biochemistry |
Departamentos de Microbiología y Ecología1, and Bioquímica y Biología Molecular2, Universitat de València, Avda Vicent Andrés Estellés s/n, 46100 Burjasssot (Valencia), Spain
Sección de Biología y Patología Celular, Centro de Investigación, Hospital la Fe3, Valencia, Spain
Author for correspondence: Daniel Gozalbo. Tel: +34 96 398 3026. Fax: +34 96 386 4299. e-mail: daniel.gozalbo{at}uv.es
The authors show that the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) of Saccharomyces cerevisiae, previously thought to be restricted to the cell interior, is also present in the cell wall. GAPDH activity, proportional to cell number and time of incubation, was detected in intact wild-type yeast cells. Intact cells of yeast strains containing insertion mutations in each of the three structural TDH genes (tdh1, tdh2 and tdh3) and double mutants (tdh1 tdh2 and tdh1 tdh3) also displayed a cell-wall-associated GAPDH activity, in the range of parental wild-type cells, although with significant differences among strains. A cell wall location of GAPDH was further confirmed in wild-type and tdh mutants by indirect immunofluorescence and flow cytometry analysis with a polyclonal antibody against S. cerevisiae GAPDH. By immunoelectron microscopy, the GAPDH protein was detected at the outer surface of the cell wall of wild-type cells, as well as in the cytoplasm. Western immunoblot analysis of cell wall extracts and cytosol showed that Tdh2 and Tdh3 polypeptides are present in the cell wall, as well as in the cytosol, of exponentially growing cells. Tdh1 is only detected in stationary-phase cells, again in both cytosol and cell wall extracts. The results incorporate the GAPDH of S. cerevisiae, encoded by TDH1–3, into the newly emerging family of multifunctional cell-wall-associated GAPDHs which retain their catalytic activity.
Keywords: insertion mutations, yeast, cell surface, glycolytic enzymes, immunodetection
Abbreviations: G3-P, glyceraldehyde 3-phosphate; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; IIF, indirect immunofluorescence; ßME, ß-mercaptoethanol; pAb anti-SC-GAPDH, polyclonal antibody against S. cerevisiae GAPDH
This article has been cited by other articles:
|
|
 |
|
  W. L. Chaffin Candida albicans Cell Wall Proteins Microbiol. Mol. Biol. Rev., September 1, 2008; 72(3): 495 - 544. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. E. Coronado, S. Mneimneh, S. L. Epstein, W.-G. Qiu, and P. N. Lipke Conserved Processes and Lineage-Specific Proteins in Fungal Cell Wall Evolution Eukaryot. Cell, December 1, 2007; 6(12): 2269 - 2277. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 F. Ferreira-da-Silva, P. J. B. Pereira, L. Gales, M. Roessle, D. I. Svergun, P. Moradas-Ferreira, and A. M. Damas The Crystal and Solution Structures of Glyceraldehyde-3-phosphate Dehydrogenase Reveal Different Quaternary Structures J. Biol. Chem., November 3, 2006; 281(44): 33433 - 33440. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. Jacob-Wilk, M. Turina, and N. K. Van Alfen Mycovirus Cryphonectria Hypovirus 1 Elements Cofractionate with trans-Golgi Network Membranes of the Fungal Host Cryphonectria parasitica. J. Virol., July 1, 2006; 80(13): 6588 - 6596. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. A. Petti and G. M. Church A network of transcriptionally coordinated functional modules in Saccharomyces cerevisiae Genome Res., September 1, 2005; 15(9): 1298 - 1306. [Abstract] [Full Text] [PDF]
|
|
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| INT J SYST EVOL MICROBIOL | MICROBIOLOGY | J GEN VIROL |
| J MED MICROBIOL | ALL SGM JOURNALS | |
