Reliable Approach for Pure Yeast Cell Wall Protein Isolation from Saccharomyces cerevisiae Yeast Cells

Abstract

Saccharomyces cerevisiae yeast is a fungus presenting a peripheral organelle called the cell wall. The cell wall protects the yeast cell from stress and provides means for communication with the surrounding environment. It has a complex molecular structure, composed of an internal part of cross-linked polysaccharides and an external part of mannoproteins. These latter are very interesting owing to their functional properties, dependent on their molecular features with massive mannosylations. Therefore, the molecular characterization of mannoproteins is a must relying on the optimal isolation and preparation of the cell wall fraction. Multiple methods are well reported for yeast cell wall isolation. The most applied one consists of yeast cell lysis by mechanical disruption. However, applying this classical approach to S288C yeast cells showed considerable contamination with noncell wall proteins, mainly comprising mitochondrial proteins. Herein, we tried to further purify the yeast cell wall preparation by two means: ultracentrifugation and Triton X-100 addition. While the first strategy showed limited outcomes in mitochondrial protein removal, the second strategy showed optimal results when Triton X-100 was added at 5%, allowing the identification of more mannoproteins and significantly enriching their amounts. This promising method could be reliably implemented on the lab scale for identification of mannoproteins and molecular characterization and industrial processes for “pure” cell wall isolation.

Proteomic Analysis of Cell Wall Proteins with Various Linkages in Fusarium graminearum

“…It is interesting that the CWPs involved in NCDB groups were enriched in GO terms related to protein synthesis, which typically occurs in the cytoplasm and endoplasmic reticulum. This could be attributed to “moonlighting” proteins with dual functionalities − or potential experimental artifacts during extraction, leading to inadvertent cytoplasmic protein contamination. , …”

“…This could be attributed to "moonlighting" proteins with dual functionalities 12−14 or potential experimental artifacts during extraction, leading to inadvertent cytoplasmic protein contamination. 9,63 Differential Expression of CWPs across Developmental Stages. CWPs in F. graminearum exhibit varied expression across developmental stages.…”

Assessment of mycotoxin sequestration efficacy in Saccharomyces cerevisiae by-products cultured in wheat bran and whey protein medium

“…In Ph disruption methods, cell fission leads to the best fragmentation of cells, which helps to effectively release proteins and other intracellular compounds. However, there is a possibility of significant contamination by proteins derived from other cytoplasmic components such as mitochondria and ribosomes 22 .…”

Importance of Non-Covalent Interactions in Yeast Cell Wall Molecular Organization

“…Considering that glucan represents most of the mass of mGC [ 2 ] and is its main structural polymer, along with a minor component, chitin [ 2 ], their remodeling is a central process that constantly occurs during growth, division and the response to changes in the environment. Thus, it can be confidently stated that the enzymes involved in remodeling are key participants in the formation and functioning of the multicomponent and polyfunctional ensemble of the CW [ 139 , 140 , 141 ].…”