De Nobel J G, Klis F M, Munnik T, Priem J, van den Ende H
Department of Molecular Cell Biology, University of Amsterdam, The Netherlands.
Yeast. 1990 Nov-Dec;6(6):483-90. doi: 10.1002/yea.320060605.
We have developed a new assay to determine relative cell wall porosity in yeasts, which is based on polycation-induced leakage of UV-absorbing compounds. Polycations with a small hydrodynamic radius as measured by gel filtration (poly-L-lysine) caused cell leakage independent of cell wall porosity whereas polycations with a large hydrodynamic radius (DEAE-dextrans) caused only limited cell leakage due to limited passage through the cell wall. This allowed the ratio between DEAE-dextran- and poly-L-lysine-induced cell leakage to be used as a measure of cell wall porosity in Saccharomyces cerevisiae, Kluyveromyces lactis and Schizosaccharomyces pombe. Using this assay, we found that the composition of the growth medium affected cell wall porosity in S. cerevisiae. In addition, we could show that cell wall porosity is limited by the number of disulphide bridges in the wall and is dependent on cell turgor. It is argued that earlier methods to estimate cell wall porosity in S. cerevisiae resulted in large underestimations.
我们开发了一种新的测定方法来确定酵母中相对细胞壁孔隙率，该方法基于聚阳离子诱导的紫外线吸收化合物泄漏。通过凝胶过滤测量的具有小流体动力学半径的聚阳离子（聚-L-赖氨酸）导致细胞泄漏，与细胞壁孔隙率无关，而具有大流体动力学半径的聚阳离子（DEAE-葡聚糖）由于穿过细胞壁的通道有限，仅导致有限的细胞泄漏。这使得DEAE-葡聚糖和聚-L-赖氨酸诱导的细胞泄漏之间的比率可用于衡量酿酒酵母、乳酸克鲁维酵母和粟酒裂殖酵母中的细胞壁孔隙率。使用该测定方法，我们发现生长培养基的组成会影响酿酒酵母中的细胞壁孔隙率。此外，我们可以证明细胞壁孔隙率受细胞壁中二硫键数量的限制，并取决于细胞膨压。有人认为，早期估计酿酒酵母中细胞壁孔隙率的方法导致了严重的低估。
