Xem trước tài liệu

Đang tải tài liệu...

Thông tin chi tiết tài liệu

Định dạng: PDF
Số trang: 19 trang
Dung lượng: 224 KB

Giới thiệu nội dung

Analysis of the vacuolar luminal proteome of Saccharomyces cerevisiae

Authors: Jean-Emmanuel Sarry, Sixue Chen, Richard P. Collum, Shun Liang, Mingsheng Peng, Albert Lang, Bianca Naumann, Florence Dzierszinski, Chao-Xing Yuan, Michael Hippler, and Philip A. Rea

Field: Proteomics, Cell Biology

Document Content: This study details a comprehensive analysis of the proteins residing within the vacuolar lumen of *Saccharomyces cerevisiae*. The research focuses on establishing a robust methodological framework for vacuole purification and subsequent proteomic profiling. By employing advanced techniques such as 2D gel electrophoresis, MALDI-TOF-MS, and LC-MS/MS, the study successfully identified a significant number of vacuolar lumen proteins. The findings provide insights into the functional roles of these proteins, many of which are involved in macromolecule degradation, homeostasis, and cellular turnover. The research also highlights the presence of proteins that may have entered the vacuole for breakdown, processing, or salvage purposes, offering a valuable resource for understanding yeast vacuolar function and for comparative studies with other eukaryotic organisms.

Detailed Table of Contents:

  • Keywords
  • Correspondence
  • Abstract
  • Introduction
  • Results and Discussion
    • Assessment of vacuolar purity and integrity
    • Resistance of vacuolar luminal proteins to proteinase K action
    • Identification of vacuolar luminal proteins
      • Canonical vacuolar and other proteases
      • Other canonical vacuolar proteins
  • Experimental procedures
    • Chemicals and yeast strain
    • Isolation of intact vacuoles
    • Preparation of vacuolar lysates
    • Fluorescence microscopy and flow cytometry
    • Vacuolar marker enzyme assays
    • Proteinase K protection assays
    • 2D gel electrophoresis
    • Gel staining, image analysis and spot intensity determinations
    • Estimation of spot molecular mass and pI
    • In-gel tryptic digestions
    • Protein identification by MS and database searching
    • Protein quantification
  • Acknowledgements
  • References
  • Supplementary material