Ramón A M, Valentín E, Maicas S, Sentandreu R
Facultat de Farmacia, Universitat de València, Avgda. Vicent Andrés Estellés, s/n, Burjassot, València, 46100, Spain.
Fungal Genet Biol. 1997 Oct;22(2):77-83. doi: 10.1006/fgbi.1997.1000.
The YWP1 gene encoding a specific mycelial cell wall protein of Yarrowia lipolytica has been cloned and expressed in Saccharomyces cerevisiae using different episomal plasmids. Because the plasmids pYAE35BB and pYAE35ES carrying the YWP1 gene (including the 5' noncoding promoter sequences) failed to express it, the YWP1 gene was cloned under the control of GAL/CYC or ACT S. cerevisiae promoters. A main band with an apparent molecular mass of 70 kDa was detected by immunoblotting in the cell wall fraction of transformants. Ywp1 processing and incorporation to the cell wall were similar in both Y. lipolytica and S. cerevisiae but not in its final localization in the cell wall. In Y. lipolytica Ywp1 is covalently bound to the cell wall (it is released only by Zymolyase digestion), whereas in S. cerevisiae it was not (it was released by boiling SDS solutions). These results suggest that the sequences involved in recognition, anchoring of a protein to the cell wall, or the catalytic activities implicated are different, at least for Ywp1, in Y. lipolytica and S. cerevisiae. Another possibility is that the target for attachment of Ywp1 is missing or cryptic in the cell wall of S. cerevisiae.
编码解脂耶氏酵母特定菌丝细胞壁蛋白的YWP1基因已被克隆，并使用不同的附加体质粒在酿酒酵母中表达。由于携带YWP1基因（包括5'非编码启动子序列）的质粒pYAE35BB和pYAE35ES未能表达该基因，因此将YWP1基因克隆到GAL/CYC或酿酒酵母ACT启动子的控制下。通过免疫印迹在转化体的细胞壁部分检测到一条表观分子量为70 kDa的主要条带。Ywp1的加工和整合到细胞壁在解脂耶氏酵母和酿酒酵母中相似，但在细胞壁中的最终定位不同。在解脂耶氏酵母中，Ywp1与细胞壁共价结合（仅通过溶菌酶消化释放），而在酿酒酵母中则不是（通过煮沸SDS溶液释放）。这些结果表明，至少对于Ywp1，参与蛋白质识别、锚定到细胞壁或相关催化活性的序列在解脂耶氏酵母和酿酒酵母中是不同的。另一种可能性是，Ywp1附着的靶点在酿酒酵母的细胞壁中缺失或隐蔽。
