Cappellaro C, Mrsa V, Tanner W
Lehrstuhl für Zellbiologie und Pflanzenphysiologie, Universität Regensburg, 93040 Regensburg, Germany.
J Bacteriol. 1998 Oct;180(19):5030-7. doi: 10.1128/JB.180.19.5030-5037.1998.
Biotinylation of intact Saccharomyces cerevisiae cells with a nonpermeant reagent (Sulfo-NHS-LC-Biotin) allowed the identification of seven cell wall proteins that were released from intact cells by dithiothreitol (DTT). By N-terminal sequencing, three of these proteins were identified as the known proteins beta-exoglucanase 1 (Exg1p), beta-endoglucanase (Bgl2p), and chitinase (Cts1p). One protein was related to the PIR protein family, whereas the remaining three (Scw3p, Scw4p, and Scw10p [for soluble cell wall proteins]) were found to be related to glucanases. Single knockouts of these three potential glucanases did not result in dramatic phenotypes. The double knockout of SCW4 and the homologous gene SCW10 resulted in slower growth, significantly increased release of proteins from intact cells by DTT, and highly decreased mating efficiency when these two genes were disrupted in both mating types. The synergistic behavior of the disruption of SCW4 and SCW10 was partly antagonized by the disruption of BGL2. The data are discussed in terms of a possible counterplay of transglucosidase and glucosidase activities.
用一种非渗透性试剂(磺基-NHS-LC-生物素)对完整的酿酒酵母细胞进行生物素化,从而鉴定出七种细胞壁蛋白,这些蛋白可通过二硫苏糖醇(DTT)从完整细胞中释放出来。通过N端测序,其中三种蛋白被鉴定为已知蛋白β-外切葡聚糖酶1(Exg1p)、β-内切葡聚糖酶(Bgl2p)和几丁质酶(Cts1p)。一种蛋白与PIR蛋白家族相关,而其余三种(Scw3p、Scw4p和Scw10p [可溶性细胞壁蛋白])被发现与葡聚糖酶相关。这三种潜在葡聚糖酶的单基因敲除并未导致显著的表型变化。SCW4和同源基因SCW10的双基因敲除导致生长缓慢,DTT从完整细胞中释放的蛋白显著增加,并且当这两个基因在两种交配型中均被破坏时,交配效率大幅降低。BGL2的破坏部分拮抗了SCW4和SCW10破坏的协同作用。本文根据转葡糖苷酶和葡糖苷酶活性可能的相互作用对这些数据进行了讨论。